Intracellular Thyroid Hormone Metabolism As a Local Regulator of Nuclear Thyroid Hormone Receptor-Mediated Impact on Vertebrate Development☆

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Biochimica et Biophysica Acta 1849 (2015) 130–141

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Biochimica et Biophysica Acta

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Review Intracellular thyroid hormone metabolism as a local regulator of nuclear thyroid hormone receptor-mediated impact on vertebrate development☆

Veerle M. Darras ⁎, Anne M. Houbrechts, Stijn L.J. Van Herck

Laboratory of Comparative Endocrinology, Department of Biology, Division of Animal Physiology and Neurobiology, KU Leuven, B-3000 Leuven, Belgium

article info abstract

Article history: Background: Thyroid hormones (THs) play an essential role in vertebrate development, acting predominantly via Received 20 February 2014 nuclear TH receptors (TRs) which are ligand-dependent transcription factors. Binding of the ligand (predominantly Received in revised form 17 April 2014 T3) induces a switch from gene activation to gene repression or vice versa. Iodothyronine deiodinases (Ds) and Accepted 7 May 2014 TH transporters are important regulators of intracellular T3 availability and therefore contribute to the control of Available online 17 May 2014 TR-dependent development. Focus: The present review discusses the possible roles of Ds and TH transporters in regulating embryonic and larval Keywords: Thyroid hormone (pre-juvenile) TR-dependent development in vertebrates. It focuses mainly on well-known model species for di- ﬁ Iodothyronine deiodinase rect and indirect vertebrate development, including zebra sh, Xenopus, chicken and mouse. Data are provided TH transporter on stage- and tissue/cell-speciﬁc changes in expression of Ds and TH transporters. This information is combined Development with functional data obtained from gain-and-loss of function studies. Vertebrate Conclusion: Knockout/knockdown of each type of D has provided strong evidence for their implication in the con- Review trol of important developmental processes and several D expression patterns and functions have been conserved throughout vertebrate evolution. Knockout/knockdown of the inactivating D3 enzyme indicates that a premature switch from unliganded to liganded TR action is often more detrimental than a delayed one. The majority of ontogenetic studies on TH transporter distribution and function have focused on brain development, showing variable impact of knockout/knockdown depending on the species. Future research in different models using conditional silencing will hopefully further improve our understanding on how TH transporters, Ds and TRs cooperate to regulate TR-mediated impact on vertebrate development. This article is part of a Special Issue entitled: Nuclear receptors in animal development. © 2014 Elsevier B.V. All rights reserved.

1. Introduction and apoptosis. Vertebrates have two types of nuclear TRs, TRα and TRβ, encoded by thra and thrb respectively. Each gene can give rise to Thyroid hormones (THs) play an essential role in vertebrate devel- different receptor isoforms via alternative splicing and each isoform opment.Asaresult,THdeﬁciency not only delays normal development, has speciﬁc functions [1,2]. However, almost all hormone binding TRs

but it also results in irreversible lifelong defects, especially in the ner- have a higher affinity for 3,5,3′-triiodothyronine (T3) as compared to vous system. In vertebrates with a clear metamorphosis, such as am- 3,5,3′,5′-tetraiodothyronine or thyroxine (T4), making T3 the predomi- phibians and many fish, TH deficiency will even stop progress from a nant hormone active at the level of the nuclear TRs in most vertebrates. larva into a juvenile and eventually result in death. THs mainly act Teleosts were recently shown to have two variants of TRβ1. While the

by binding to nuclear TH receptors (TRs) that control transcription of short variant predominantly binds T3, the long variant binds with high TH-responsive genes. Many of them are important regulators of devel- afﬁnity to 3,5-diiodothyronine (3,5-T2) and both hormones are physio- opmental processes, including cell division, differentiation, migration logically relevant in regulating gene expression [3,4].

Abbreviations: D1, D2, D3, iodothyronine deiodinase types 1, 2, and 3; KD, knockdown; 2. Need for local regulation of TH availability KO, knockout; LAT1, LAT2, L-type amino acid transporters 1 and 2; MCT8, MCT10, monocarboxylate transporters 8 and 10; OATP1C1, organic anion transporting polypeptide 10; The only cells capable of producing THs are the follicular cells of the TH, thyroid hormone; TRα,TRβ, thyroid hormone receptors α and β thyroid gland. This gland mainly releases T4 while T3 is released in ☆ This article is part of a Special Issue entitled: Nuclear receptors in animal development. minor amounts. During the early stages of development, embryos are ⁎ Corresponding author at: Laboratory of Comparative Endocrinology, Naamsestraat 61 P.O. Box 2464, B-3000 Leuven, Belgium. Tel.: +32 16 323 985; fax: +32 16 324 262. not yet able to synthesize their own THs. They therefore rely on mater- E-mail address: [email protected] (V.M. Darras). nal THs that are deposited in the yolk or supplied via the placenta in the

http://dx.doi.org/10.1016/j.bbagrm.2014.05.004 1874-9399/© 2014 Elsevier B.V. All rights reserved. V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141 131 case of mammals. As development progresses and the embryonic thy- also a low Km enzyme that catalyses only inner ring deiodination roid gland starts functioning, the relative contribution of maternal THs (IRD) and has T3 as preferred substrate. Its main function is the conver- gradually declines. Maternal and embryonic THs are transported via sion of T3 as well as T4 into iodothyronines that do not bind to nuclear the circulation throughout the body to reach the different tissues and TRs. In general, a given cell type expresses either one or two types of the amount of hormone in circulation changes according to a species- Ds but the number and type may change several times during ontogeny. specific ontogenetic pattern. However, embryonic or larval tissues do Development in vertebrates can be either direct or indirect. In not all develop in the same way nor at the same rate. Therefore different mammals, birds and reptiles the process of birth (following internal tissues/cell types need several mechanisms to modulate the impact of development) or hatching (following external development) marks circulating THs according to their specific needs at a given time. the direct transition from the embryo to the juvenile, immature adult One common mechanism to regulate local hormone action is by form. Amphibians and fish have an indirect development where hatch- changing the expression of hormone receptors. The expression of differ- ing is followed by a distinct larval phase separating the embryo and ent TRs indeed changes throughout development in a tissue-specific the juvenile. This review focuses on some well-studied species with way [5–7]. However, TRs not only function when bound to their ligand indirect/direct development to illustrate the impact of Ds on each of since unliganded TRs also bind to TH-response elements (TREs) in the the pre-juvenile stages and on the sequential transition from one promotor region of TH-responsive genes. In case of a positive TRE, bind- stage to the other. Whenever possible, we try to combine information ing of the ligand can acutely reverse TR impact from repression to stim- on D ontogenetic expression patterns with information gained from ulation of gene transcription while in case of a negative TRE the opposite gain-and-loss of function studies. may occur [2,8,9]. Changes in intracellular ligand availability are therefore an essential level to regulate TR-mediated action. Intracellular 3.1. Indirect development in fish and amphibians iodothyronine deiodinases are important in this process since these enzymes can locally produce or degrade T3 as well as 3,5-T2 [4].THtrans- 3.1.1. Zebrafish embryos porters located in the plasma membrane are important as well, since Embryonic D expression has been studied in detail in zebrafish they control influx of THs into the cell as well as TH efflux (Fig. 1). The (Danio rerio), one of the major vertebrate models for developmental bi- present review focuses on the role of changes in iodothyronine ology. Zebrafish embryonic development takes about 3 days and their deiodinases and TH transporters in vertebrate development, while thyroid gland starts hormone secretion around hatching [15]. Due to a other reviews in this special issue on Nuclear Receptors in Animal whole genome duplication in the early evolution of ray-finned fishes, Development focus more specifically on the changes at the level of the zebrafish and several other teleost fish have two dio3 paralogues [16, TRs. 17], similar to the presence of two thra paralogues [18]. Maternal mRNA for all three types of Ds is present in the egg and by 3. Iodothyronine deiodinases and vertebrate development the end of the maternal–zygotic transition around 8 h post fertilisation (8 hpf) embryonic transcripts can be detected and quantified. While Vertebrates possess three types of iodothyronine deiodinases (D1, whole body dio1 and dio3 mRNA levels increase during the first day of D2, D3) encoded by dio1, dio2 and dio3 respectively [10–12].D1isa embryogenesis, dio2 mRNA levels only rise substantially on the third multifunctional enzyme that can cleave iodine from the outer as well day, around hatching concomitant with the start of embryonic TH secre- as the inner ring of an iodothyronine. It is a high Km enzyme that can tion ([17,19]; and own unpublished results) (Fig. 2). In situ hybridiza- convert T4 into T3 and as such contributes to circulating T3 levels. Its pre- tion (ISH) results on the tissue-specific expression pattern in zebrafish ferred substrate is however reverse T3 (rT3) and studies in D1-deficient embryos have been reviewed recently [20]. At 12 hpf the dio1 and mice suggest that D1 functions primarily as a scavenging enzyme, im- dio2 signals are colocalized in the head region while the dio3 signal is portant in iodine recycling from inactive and lesser iodothyronines found in the kidney. From 24 hpf onwards dio2 is also expressed in [13,14]. D2 only catalyses outer ring deiodination (ORD); it is a low the retina and the adenohypophysis. At 48 hpf dio2 also appears in the

Km enzyme with T4 as major substrate. It is important for local T3 pro- spinal cord and dio1 canbedetectedintheliver[21–23]. duction in several tissues but also contributes to circulating T3,especial- Thanks to the relatively easy method of antisense morpholino injec- ly in ﬁsh and amphibians. It is also a likely candidate to convert T3 into tion [24] loss-of-function studies in zebraﬁsh embryos have been possi- 3,5-T2, which binds to the long variant of TRβ1inteleosts[3].D3is ble for all three Ds. Knockdown (KD) of D2 clearly delays development

Fig. 1. Intracellular T3 availability and action. Specific TH transporters facilitate the influx as well as the efflux of T4 and T3 (and some other iodothyronines). Within the cell, different deiodinases (Ds) activate or inactivate THs, thereby regulating T3 availability. T3 translocates to the nucleus where it binds to TH receptors (TRs). TRs form heterodimers (predominantly with the retinoic X receptor, RXR) and bind to TH response elements (TREs) in the promoter region of TH-responsive genes. The figure represents the situation for positively regulated genes where unliganded TRs repress gene transcription, while binding of T3 induces a switch from repression to activation of gene transcription. The opposite occurs for negatively regulated genes. 132 V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141

research group, showing impaired caudal cranial neural crest development, and as a result, inhibited development of the posterior pharyn- geal arches, ceratohyal cartilage and iris stroma [27].

3.1.2. Xenopus embryos Frog development from fertilisation up to completion of metamorphosis is generally divided in a number of consecutive stages based on morphological features. For Xenopus they have been described in detail by Nieuwkoop and Faber (NF stages 1 to 66) [28]. In the African clawed frog (Xenopus laevis) mRNAs of all three Ds are present throughout embryogenesis, showing that in contrast to what was long accepted, the dio1 gene is also present and transcribed in amphibians [29,30]. dio1 mRNA is the predominant maternal D transcript present in eggs but levels decline rapidly. Embryonic dio1 mRNA expression sharply increases around NF45 when the embryonic thyroid gland starts functioning. Embryonic dio2 and dio3 mRNA levels increase earlier, starting respectively around NF41 and NF37 [30]. ISH during late embryogenesis (NF30–40) shows strong expression of the three dio genes in the head region and axial structures but the expression patterns do not fully overlap. dio1 expression is strong in the notochord and branchial arches, dio2 expression is mainly found around the eye and the neural tube and dio3 expression is mainly present around the branchial arch areas and around the eye. It could also be shown that dio2 and dio3 mRNA are translated into functional proteins (tested at NF37) while the contribution of D1 to deiodination seems to be minor [29]. Fig. 2. Ontogenetic profile of mRNA levels for TH receptors (thra and thrb) and deiodinases ISH studies in the western clawed frog (Xenopus/Silurana tropicalis) (dio2, dio3a and dio3b) in whole body extracts of zebrafish (Danio rerio)embryosandearly larvae up to 7 days post fertilisation (dpf). Data are presented as fold change relative to the show expression of dio3 but not dio2 in the gastrula. From neurulation lowest level measured for each separate gene. Pictures representing different develop- until the embryo–larva transition (stages NF22 to NF36), a dio2 signal is mental stages are shown below the time-axis. Embryos normally hatch between 50 and present in the notochord, brain, retina and the ventral blood-forming 70 h post fertilisation, indicated by the black bar. Both thrb and dio3b levels increase sharp- region and dio3 mRNA is consecutively detectable in the ventral ly on the first 2 days post fertilisation while dio3a and dio2 expression increase sharply around hatching. dio1 mRNA levels are not included in the figure for clarity. They double blood-forming region, the retina, the notochord, the pro-nephros and by the end of 1 dpf and then remain stable up to 3 dpf (own unpublished results). the otic vesicle [31]. Taken together, the results available from both Data are taken from [19,41,136]. Xenopus species indicate that the expression patterns of Ds in embryonic tissues such as brain are highly dynamic [31,32]. Since TRs are also present in Xenopus embryos from the start of development [33,34], deiodinases may serve to locally regulate the balance between activa- as demonstrated by morphological parameters such as otic vesicle tion and inactivation of maternally derived THs and hence the balance length, head-to-trunk angle and pigmentation index. It also significantly between TR-mediated gene repression and activation [35]. decreases thrb expression [25]. KD of D1 does not have obvious effects Unfortunately, what we know about the role of Ds in amphibian em- on embryonic development, but combined KD of D1 and D2, blocking bryonic development is so far mostly based on correlations. Functional all T4-activating pathways, aggravates the phenotype observed in D2 proof awaits results from embryos lacking or overexpressing specific morphants. Most D1D2 morphants show delayed/aberrant eye devel- Ds, an approach that is technically possible and will hopefully provide opment (own unpublished results) and some embryos do not develop new data in the near future. an inner ear and/or have severely curved tails [26]. This shows that while D1 may not be essential for embryonic development in euthyroid 3.1.3. Embryo-to-larva transition in zebrafish zebrafish, it may be important in conditions where T3 availability is re- One of the major events accompanying the embryo-to-larva transi- duced due to impaired D2 activity. This illustrates the importance of pe- tion in zebrafish is inflation of the swim bladder, important for active ripheral T3 production and indicates that liganded TR signalling is swimming behaviour and feeding. Zebrafish D3b morphants show a se- required for normal progression of several essential developmental vere delay in swim bladder inflation, although timing of the formation processes. of the swim bladder anlage seems normal. One hypothesis is that the

Recently we also published results for KD of both T3-inactivating D3 delayed inflation is due to the substantially reduced motility observed enzymes, the long known D3b and the more recently identified D3a in the prehatch embryos and the larvae, reducing their ability to gulp [17]. Some of the morphological changes observed before in D1D2 air at the water surface [17]. THs could also more directly interfere morphants are also present in D3b morphant embryos, including an with swim bladder development, possibly by interaction with the overall delay in development and defects at the level of the eye [17]. hedgehog and/or the canonical wnt signalling pathways. Both pathways This suggests that several processes in zebrafish embryonic develop- are known to be important regulators of zebrafish swim bladder devel- ment are highly dependent on the exact balance between unliganded opment [36,37] and both have already been linked to THs in other pro- and liganded TR signalling. Analysis of the transcriptome around hatch- cesses [38,39]. Changes in D2 and/or D3 activity induced by hedgehog, ing (72 hpf) shows that some changes in gene expression are similar in as known to appear for instance in differentiating chicken chondrocytes D1D2 and D3b morphants, but there are also many divergent responses and in proliferating mammalian keratinocytes [40], may add an addi- (own unpublished results). We also knocked down D3a and found that tional level in the interaction of these three signalling pathways and the phenotype is similar but significantly milder than following D3b KD. hence affect swim bladder development in zebrafish, although this hy- This could be matched to enzyme activity studies showing that KD of pothesis remains to be tested. D3b almost completely abolishes D3 activity in homogenates of 72 hpf We found a similar but milder defect in swim bladder inflation in embryos while D3a KD reduces activity by only 22–26% [17]. Results D3a morphants but also in D1D2 morphants. While the latter may on morpholino based KD of D3b are also available from another seem surprising, it corresponds with other studies showing that both V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141 133

TH supplementation and induction of hypothyroidism interferes with zebraﬁsh swim bladder inﬂation [41]. It might just be another example of how important the exact balance between TR-mediated gene activation and repression is for development to progress normally.

3.1.4. Larva-to-juvenile transition in frogs Amphibian metamorphosis is probably the best known example of a developmental process controlled by THs. That is probably also the reason why much of the pioneering work on the role of peripheral TH deiodination in development started in this model. It was suggested already in 1962 that enzymes metabolizing THs might be important regulators of metamorphosis [42]. Following later studies on ORD

(T3-producing) and IRD (T3-degrading) activity in premetamorphic and metamorphosing tadpoles, Galton concluded in 1988 that next to the number of TRs, the balance between ORD and IRD activities in a given target cell regulates the effectiveness of circulating THs during metamorphosis [43]. The metamorphosing tadpole was also the ﬁrst species where a D3 enzyme was cloned following its identiﬁcation as an early response gene in TH-induced tail resorption [44,45].

A detailed analysis of D2 and D3 activity and mRNA expression Fig. 3. Changes in D2 and D3 activity in different tissues of metamorphosing bullfrog (Rana during bullfrog (Rana catesbeiana) development subsequently demon- catesbeiana). Arrows indicate respectively the start of rapid hindlimb growth in early strated that the timing of major metamorphic changes and highest prometamorphosis (1), the eruption of forelimbs at the start of metamorphic climax (2) dio2 mRNA levels/D2 activity coincide in many tissues, including limbs, and the start of tail shrinkage (3). D2 and D3 activity peak simultaneously in hindlimb during growth and increase together in forelimb around emergence. In tail, D2 activity in- tail, intestine, skin and eye. Blocking D2 activity with the deiodinase increases sharply when tail shrinking starts while D3 activity peaks earlier. hibitor iopanoic acid (IOP) clearly retards metamorphic events in these Developmental stages are indicated according to Taylor and Kollros [137] (TK-stages V to tissues [46]. The link between dio2 expression and metamorphic changes XXIV) and representative images for some metamorphic changes are taken from [138]. was also made in X. laevis by studying TH-stimulated cell division. Cells Data are taken from [46]. in brain, spinal cord and limb buds that proliferate in response to low circulating TH levels during the early phase of metamorphosis, constitu- 3.1.5. Larva-to-juvenile transition in teleost fish fi tively express high levels of dio2 and their response to T4 can be The possible role of THs in larva-to-juvenile transition in sh has blocked by IOP treatment [47]. Both cell division and dio2 expression been studied mainly in species where this transition is accompanied in these regions decrease at metamorphic climax, when dio2 expression by clear changes in morphology and habitat. One example is flatfish, increases in response to high circulating TH levels in tissues such as in- where metamorphosis transforms a bilaterally symmetric pelagic larva testine and tail that do not proliferate but remodel or disappear [47]. into an asymmetric benthic juvenile. TH secretion and TR expression The link between the expression pattern of D3 and metamorphosis increase during spontaneous metamorphosis and the process is ac- seems to be more complex. In several, although not all bullfrog tissues celerated by TH supplementation [52–55]. In Senegalese sole (Solea the timing of greatest metamorphic changes and high dio2 mRNA/D2 senegalensis) measurements in whole body extracts show that both activity coincides also with increased expression/activity of the T3- dio2 mRNA and D2 activity keep increasing from the start to the end of degrading enzyme D3 [46] (Fig. 3). In contrast, expression of D3 in metamorphosis (Fig. 4). dio3 mRNA levels show a peak at metamorphic X. laevis correlates with a tissue having a reduced response to TH and climax, but D3 activity decreases from the start to the end of metamor- D3 is upregulated prior to metamorphic change or just before the phosis [54]. D2 expression is also high in metamorphosing turbot completion of a tissue's response to TH [48].Thesefindings may be (Scophtalmus maximus) [52]. Changes in D expression also occur in the complementary, illustrating the multiple roles of D3 in tadpole tissues: larva-to-juvenile transition of teleost species that undergo less dramatic (1) reducing TH responsiveness and allowing the action of unliganded changes, such as sea bream (Sparus aurata). Expression of D1 and D2

TRs prior to metamorphic changes, (2) fine-tuning intracellular T3 levels starts to increase at the start of metamorphosis, peak at climax together in combination with D2 at the moment of metamorphic changes, and with TR expression and decline again towards the end of metamorpho- (3) stopping the liganded TR-induced gene expression programme at sis. D3 expression, however, does not change during the process [56]. the end of metamorphic changes [46,49]. The increase in ORD and decrease in IRD activity together with increased The crucial impact of D3 on metamorphosis was demonstrated in TR availability at the start of metamorphosis suggests that liganded TR transgenic X. laevis tadpoles overexpressing D3. Gill and tail resorption signalling through binding of T3, and possibly 3,5-T2, is a driving force were clearly delayed and most of the animals arrested metamorphosis in teleost metamorphosis. and died [50]. The same research group also showed the impact of D3 Results on whole body expression were recently complemented in the eye. Prior to metamorphosis, the retina grows symmetrically. As by tissue-specific ISH data in metamorphosing flounder (Paralichthys TH levels start to rise with the onset of metamorphosis, the morphology olivaceus) [16]. dio1 expression is localized in the liver and is restricted of the eyes starts changing. Cell proliferation in the ventral ciliary mar- in time from prometamorphosis to early climax; it is thought to be impor- ginal zone (CMZ) increases and ipsilateral projections to the brain are tant for supply of T3 to the blood. dio2 is expressed in eye, tectum and formed, concomitant with the change in head shape that allows some skeletal muscles throughout metamorphosis. dio3 is expressed from overlap in visual fields. In contrast to the proliferating cells of the ventral premetamorphosis until early climax in several tissues, including skeletal CMZ, cells in the dorsal CMZ express high levels of D3 and fail to respond muscle and the developing stomach [16]. As is the case in amphibians, to the increase in circulating THs, resulting in asymmetrical retinal these expression patterns suggest that the three types of Ds are specific growth [51]. In transgenic tadpoles overexpressing D3 throughout the and important regulators of larva-to-juvenile transition in teleosts. body, cell division in the ventral CMZ was reduced and ipsilateral projections to the brain were not formed [49]. No additional data on trans- 3.2. Direct development in reptiles, birds and mammals genic tadpoles lacking or overexpressing Ds were published more recently which is unfortunate, since amphibian metamorphosis remains Birds and most reptiles have an external development where the an excellent model to study D involvement in development. hatching process marks the transition from the embryonic to the 134 V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141

present, followed by D1, while D2 expression seems to be restricted mainly to brain [64]. Studies with the more sensitive RT-PCR and ISH techniques are only available for some embryonic tissues. Our own research group focused on brain development. mRNA for all three Ds is present in brain from the first stage studied, namely 4-day-old embryos (E4), but unlike DIO2 and DIO3, DIO1 mRNA does not seem to yield an active enzyme. DIO2 mRNA and D2 activity gradually increase in all major brain regions from E4 to E12 while the patterns for DIO3/D3 are more variable amongst brain regions [65]. This fits with ISH data from E3 to E10 showing that DIO2 is expressed throughout the brain, predominantly associated with blood vessels, while it is absent in the choroid plexus at this time [66,67]. In contrast, high DIO3 expression is specifically localized in the choroid plexus and in a limited number of cell populations, including several brain regions involved in the development of the visual system [66]. Dynamic and cell-specific changes also occur at the protein level, as demonstrated for all three Ds by immunohistochemistry (IHC) in cerebellum in the days around hatching. For instance, E18 Purkinje cells stain weakly positive for D2 but not for D3 while in 1-day-old chicks they stain strongly positive for D3 only. This shift is accompanied by a strong decrease in the presence of reelin and Dab-1 proteins, both prod- ucts of TH-responsive genes [68]. Another example is found at the level of the choroid plexus. While DIO3 mRNA and D3 protein expression dominate in early embryonic stages, there is a transient increase in DIO2/D2 expression in the days just prior to hatching ([69] and own un-

published results). This could be an additional source for T3 entry into the brain in the last days of embryonic development, when major maturation processes take place. Since this coincides with a period of fi Fig. 4. Ontogenetic pro le of mRNA levels for TH receptors (thraa2 and thrb2)and increased expression of THRB, especially in cerebellum, signalling via deiodinases (dio2, dio3) as well as deiodinase activities (D2 and D3) in whole body ex- β tracts of metamorphosing flatfish (Solea senegalensis). Data are presented as fold change liganded TR seems to be important for perinatal glial and neuronal relative to the lowest level measured for each separate gene. Metamorphosis starts around maturation [70]. 12 days post fertilisation (dpf) and is completed by 21 dpf. Expression of both TH receptor Developing chicken eye and ear show a highly dynamic expression isoforms peaks in the second half of metamorphosis. dio2 expression and D2 activity con- pattern of D2 and D3, comparable to what has been observed in mice tinue to increase up to the end of metamorphosis while D3 activity gradually declines from early to late metamorphosis despite a transient peak in dio3 expression. (see below). While DIO3 mRNA is already present in chicken retina at Representative images of sole developmental stages have been taken from [139].Dataare E3, DIO2 only appears around E6 in the outer neuroblastic layer. At taken from [54]. E8, DIO2 is expressed in the outer nuclear layer, presumably in developing photoreceptors, while DIO3 is mostly confined to the inner nuclear juvenile stage. While newly hatched reptiles are in general quite self- layer, presumably associated with retinal progenitor cells (Fig. 5). By supporting, this is not the case in many birds. Songbird hatchlings are E10, DIO2 expression is restricted to the area adjacent to the lens poorly developed and completely dependent on parental care for sur- [66,71].ThepeakinDIO2 expression in the immature photoreceptor vival. Songbirds and several other bird species are called altricial, in con- layer coincides with a peak in THRB expression suggesting that D2- trast to precocial birds like chickens and ducks where hatchlings are mediated TRβ activity is necessary for specific stages in photoreceptor mobile and capable of finding food. This difference in precocial versus differentiation [71]. Ds probably also have a role in development of altricial development clearly coincides with differences in timing of the cornea which gradually becomes transparent from E12 to E20. the start and maturation of TH secretion and peripheral TH metabolism, This transformation coincides with a gradual increase in expression of suggesting there is a link [57,58]. For mammals too, we can identify al- THRB and DIO3 while DIO2 expression peaks at the time the process tricial species like rodents and precocial species like ungulates although starts and declines thereafter. THRA and DIO1 expression on the other many species, including humans, should be classified somewhere in be- hand remain constant [72]. In the inner ear, DIO2 expression starts at tween. Typical model species for both types of development are the pre- E6 and gradually increases during development while no DIO3 expres- cocial chicken and the altricial mouse. sion has been detected by ISH [66]. For peripheral tissues, the cellular distribution of D1 and D3 proteins has been described in liver and kidney [73]. In the last week of embry- 3.2.1. Chicken embryo onic development, high levels of both Ds are present in a thin layer of Chicken embryonic development takes 3 weeks; the thyroid gland hepatocytes around the blood vessels. In kidney, D1 and D3 are found starts hormone secretion after approximately 7 days of incubation in several cell types, including the tubular epithelial cells. This could

(E7) and is fully functional around E12 [59]. Studies on TH deiodination mean that a tight regulation of T3 is necessary for correct tubular func- in avian embryos started in the 1980s by in vitro characterization of tion but it could also reflect a function of these cells in the recuperation ORD and IRD activities in chicken embryonic liver and heart [60–62]. of iodine from the renal filtrate. A study in embryonic chick tibial It was shown by several groups that a decrease in hepatic IRD activity growth plates has shown that D2 is expressed in chondrocytes and in the last days prior to hatching is a major factor contributing to the that there is a direct interaction between the hedgehog and TH- sharp increase in circulating T3 that precedes hatching [60,61,63].Fol- signalling pathway at that level [40]. A local increase in Indian hedgehog lowing the identification and cloning of the three D types in chicken, leads to increased D2 degradation via the ubiquitination pathway, their expression pattern was studied during the third and last week of resulting in a reduction of local T3 production. This leads to an induction embryonic development using Northern blot and in vitro kinetic tests. of parathyroid hormone-related peptide, which in turn will control the These techniques showed that D3 (mRNA and activity) is most widely transition from chondrocyte proliferation to differentiation [74]. V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141 135

and the transition from the embryonic to the posthatch stage, studies with DIO gene silencing or overexpression would be very helpful in elucidating the speciﬁc role of each one of the Ds. In vivo KD by RNAi- induced silencing is a possible approach. It has already been proven suc- cessful for KD of D2 expression in chicken retina. Unfortunately, the authors only used it as a proof of principle and did not discuss the resulting effects on retinal development [83]. This would have been very interesting for comparison with the eye phenotype observed in D2 knockout mice (see below).

3.2.2. Prenatal and neonatal mouse Rat and mice were amongst the first vertebrates where peripheral deiodination was investigated, starting in the 1950s [84], but for long the studies mainly focused on postnatal life stages. This might be linked to the intra-uterine development, making experimental manipulations in embryonic/foetal rodents less easy, but also to the fact that the thyroid gland in rodents develops late. Foetal TH secretion only starts in the third and last week of intra-uterine development and the thyroid axis only matures by the end of the first postnatal week [85,86]. This is linked to their altricial development and many developmental processes that occur prior to birth in more precocial mammals can be studied postnatally in rodents. With the ever increasing possibilities of complete as well as conditional deiodinase knockout (DKO), the number of developmental studies in mice has increased exponentially over the last 10 years and several of them now also include prenatal stages. One factor complicating the study of Ds and DKO in mice is that Dio3 is an imprinted gene [87]. Genomic imprinting is an epigenetic phe- nomenon in mammals that affects a small number of genes and results in the preferential expression of one of both alleles, depending on the parental origin [88]. The imprinted gene cluster containing Dio3 is normally expressed from the paternally inherited chromosome, but the maternal Dio3 allele does not seem to be completely silenced. Dio3 imprinting is not complete in the placenta and its expression can be bi- allelic or even maternally inherited in certain areas of the brain [89]. Fig. 5. Distribution pattern of DIO2, DIO3 and MCT8 mRNA shown by ISH in the retina Maternal transfer of THs to the developing embryo/foetus occurs via of 8-day-old chicken embryos. Positive layers (dark blue) are identified in white letters, the placenta. It was originally thought that rodent prenatal tissues con- other layers are identified in black. The image for D2 is from a peripheral area in the retina (near the lens) where D2 expression is highest. The images for D3 and MCT8 are from the tain only low levels of T3 because foetal TH secretion starts late and the central retina (near the optic nerve). Scale bars represent 100 μm. GCL: retinal ganglion high IRD activity in placenta limits maternal T3 transfer [90].However, cell layer, INL: inner nuclear layer, OFL: optic fibre layer, ONL: outer nuclear layer, RPE: the distribution pattern of Ds allows the foetus to increase or decrease retinal pigment epithelium. T3 availability in a tissue-specific way and also to compensate at least partially for changes in maternal TH supply [91]. In rat foetal liver, kid- While the above interaction with Indian hedgehog is an example of a ney, intestine, lung and eye, the principal TH-activating enzyme is D1, localized influence on Ds and T3 availability, some hormones from out- while D2 is the principal TH-activating enzyme present in brain and side the thyroidal axis are changing D expression with consequences for the only D present in brown adipose tissue. Foetal skin expresses all circulating T3 levels throughout the body. A good example is the in- three Ds. D3 activity levels are high in many foetal tissues, including crease in circulating T3 prior to hatching. Both growth hormone and liver, brain, intestine and skeletal muscle. In general, D3 levels tend to corticosterone (or its synthetic analogue dexamethasone) acutely shut decrease towards birth while D1 and D2 levels increase [92,93]. down DIO3 gene expression in embryonic chicken liver [75].Liveris Our understanding of the role of specific Ds in the development and responsible for a substantial part of peripheral T3 production and degra- maturation of different tissues has greatly improved by the study of dation, and hepatic D3 activity is very high until some days before DKO mice. These models prove, somewhat surprisingly, that foetal sur- hatching [63]. Injection of growth hormone or glucocorticoids at that vival is possible without expression of any activating D [94]. The same is stage rapidly decreases hepatic D3 activity and hence increases plasma true for D3 deficiency, although this is accompanied by partial neonatal

T3 [76,77]. The rise in plasma growth hormone and corticosterone lethality [87]. More detailed studies of the phenotype of D1KO mice starting a few days before hatching therefore seems to be a determining show marked changes in the metabolism and excretion of THs, but pro- factor in the rise in plasma T3, necessary for hatching [63,78].Glucocor- vide little evidence for a major role of D1 in mouse development [14,92]. ticoids also have an effect in embryonic brain where they decrease DIO3 In contrast, D2KO and especially D3KO mice show developmental ab- and increase DIO2 gene expression [75] resulting in local changes in T4 normalities in several tissues, some of them leading to life-long deﬁcits. and T3 availability [79]. This regulatory role of glucocorticoids may be D2KO mice have normal plasma levels of T3 but increased levels of important in vertebrate development in general, since they induce sim- T4 and thyrotropine (TSH) as a result of pituitary resistance to T4 [95]. ilar changes in D expression in larval (neotenic) axolotls (Ambystoma D3KO mice show a complicated pattern of thyroid function. They mexicanum) and at least partially also in bullfrog tadpoles and rat show a several-fold increase in plasma T3 compared to wild type both foetuses [80,81]. The effect is however stage dependent, and glucocorti- in utero and in the neonatal period, due to a reduced T3 clearance rate coid treatment tends to decrease plasma T3 in posthatch/postnatal birds [96]. However, from postnatal day 15 up to adulthood they demonstrate and mammals both by central and peripheral actions [76,80,82]. central hypothyroidism with low plasma T3 and T4 levels, suggesting Although the above examples of interaction with Ds already show that the neonatal thyrotoxicosis has severely disturbed the maturation that these enzymes help in regulating chicken embryonic development of the hypothalamo-pituitary-thyroid axis [96]. 136 V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141

Although TH levels in circulation of D2KO mice are sufficient, local T3 days. The number of rods, which mediate dim light vision and do not ex- signalling is reduced in several tissues. D2 deficiency lowers the expres- press TRβ2, remain normal [110]. sion of TH-responsive genes, including Pgc-1α and Ucp1,inembryonic Some studies have already shown changes in brain development brown adipose tissue. This results in impaired differentiation and oxida- mainly at the level of gene expression, although a lot remains to be ex- tive capacity of brown adipocytes, and results in permanent defects in plored. In the first days after birth, T3 content is significantly increased adaptive thermogenesis [97]. Local T3 production by D2 is also impor- in D3KO brain, resulting in increased expression of TH-responsive tant for muscle development. Dio2 is expressed in proliferating muscle genes such as Hairless and RC3 [96]. By two weeks after birth, when pe- progenitor cells and increases markedly during differentiation as a re- ripheral TH levels have dropped, the picture seems to be more complex. sult of an increase in the transcription factor FoxO3. Myotube formation Studies with a transgenic mouse model (FINDT3), expressing β galacto- is blocked in myoblasts of D2KO mice and the normal myogenic pro- sidase as a readout of local T3 availability, have shown that by that time gramme, including induction of MyoD, myogenin and MHC, is inhibited. some brain regions still have increased T3 levels, while T3 levels in some This affects muscle development as well as muscle regeneration after other regions have already decreased [111]. This indicates that D3 has injury [98]. It has been found recently that the increase in D2 expression an important and region-specificroleinregulatingT3 signalling in the in differentiating myoblasts involves an epigenetic control mechanism developing brain. T3 content in the neonatal D2KO brain is markedly re- via the lysine-specific demethylase enzyme LSD-1 which relieves the duced. Nevertheless, expression of several TH-responsive genes is not or repressive marks on the Dio2 promoter. This enzyme also leads to only mildly affected, suggesting that compensatory mechanisms must a downregulation of D3 due to removal of activation marks on the occur, limiting the impact of D2 deficiency [105]. A recent study on

Dio3 promoter [99]. Both processes contribute to increased local T3 the expression of cerebral cortex genes in 4-week-old mice indicates availability. that the impact of D2 and D3 deficiency is higher on negatively regulat- The bones of adult D2KO mice have reduced toughness and are ed TH-responsive genes than on positively regulated ones [112].Finally, brittle, due to increased skeletal mineralization. However, investigation a recent study on cerebellum shows that D2 activity is very low in the of the skeleton of D2KO mice at birth reveals no abnormalities, and en- late foetal stages and at birth but increases during the first postnatal dochondral ossification in D2KO mice is normal [100].Thisisconsistent week, while D3 activity is relatively high prenatally and starts to de- with the finding that, in contrast to chicken, mouse growth plate crease 3 days after birth [106]. As a consequence of aberrant timing of chondrocytes do not seem to possess D2 activity, while the foetal T3 signalling, D3KO mice show several abnormalities in cerebellar de- growth plate expresses high levels of Dio3 mRNA that decrease sub- velopment, including reduced foliation, accelerated disappearance of stantially towards birth [101,102]. The adult brittle bone phenotype the external germinal layer and premature expansion of the molecular therefore does not seem to be a consequence of abnormal skeletal de- layer. Combined deletion of the TRα1 substantially corrects the cerebel- velopment [100]. lar phenotype but for instance not the deafness of D3KO mice where D2KO and D3KO mice also show changes in carbohydrate and lipid TRβ signalling is important. This illustrates that the combined expres- metabolism. Dio3 is highly expressed in embryonic as well as adult pan- sion of D3 and specific receptor isoforms is controlling development of creatic islets, predominantly in β cells, while Dio1 and Dio2 mRNA levels different tissues [106]. are very low. Neonatal D3KO mice have a reduced β cell mass, reduced insulin production and impaired expression of several key β cell genes. This developmental defect persists throughout life and leads to dis- 4. TH transporters and vertebrate development turbed glucose homeostasis [103]. Glucose homeostasis is also disturbed in D2KO mice which show insulin resistance [104].Inaddition, Although it was accepted for long that THs enter the cell by simple they have an increased weight gain compared to wild type when placed diffusion, it is now clear that their passage through the plasma mem- on a high fat diet, a characteristic that might be linked to impaired brane depends on specific transmembrane transporter proteins. Thy- brown adipose tissue development and function as mentioned above. roid hormone transporters belong to different transporter families: Although D2KO and D3KO mice do not show a severe neurological the Na+/taurocholate cotransporting polypeptide (NTCP) family, the phenotype at first sight, more detailed behavioural studies start to re- Na+ independent organic anion transporting polypeptide (OATP) veal smaller defects, e.g. the impaired ability to invert and descend a family, the L-type amino acid transporter (LAT) family and the mono- vertical pole [105,106]. The possible link of D deficiency with the struc- carboxylate transporter (MCT) family [113]. The transporters with tural development of ear, eye and brain has been studied to a large ex- the highest affinity for THs are MCT8 (encoded by slc16a2), MCT10 tent during the first postnatal weeks, consistent with the relatively (encoded by slc16a10) and OATP1C1 (encoded by slco1c1). The general late maturation of the rodent brain and sensory systems. Expression of picture coming from the few species studied is that MCT8 and MCT10

Dio3 is high in the immature prenatal cochlea and decreases in the efficiently transport T4 and T3. Rat (but not human) MCT8 also efficient- first days after birth. In contrast, Dio2 expression is very low in prenatal ly transports rT3 and 3,3′-T2. OATP1C1 shows preferential transport of cochlea but increases sharply in the first postnatal week, prior to the T4 and rT3 [114]. Other transporters that also seem to be of physiological onset of hearing in the second week [107].D2deficiency delays cochlear importance are LAT1 (encoded by slc7a5) and LAT2 (encoded by slc7a8), development while D3 deficiency accelerates the process. However, but they transport T3 and T4 with a much lower affinity [113].Itisim- both conditions lead to impaired development of the sensory epitheli- portant to take into account that all these transporters do regulate not um and permanent deafness [108,109]. The cellular expression pattern only TH entry into the cell but also TH efflux, allowing passage of acti- of D3 partly overlaps with that of TRβ, the receptor that is primarily vated hormone from one cell type to another [114]. responsible for auditory development [109]. These studies are a clear A clear proof for the essential role of TH transporters in development example of how essential local changes in the balance between TH acti- was the discovery that the human Allan–Herndon–Dudley syndrome, vation and inactivation are, but also of the fact that these changes have characterized by hypotonia, weakness, reduced muscle mass, severe to occur within a critical time window to ensure normal development. mental retardation and several other neurological symptoms, is linked

Correct T3 signalling is also important for eye development. D3 activ- to mutation of the MCT8 gene [115,116]. Studies in MCT8KO mice ity levels are high in prenatal mouse retina while little or no D1 or D2 were started soon after this discovery, but the neurological phenotype activity can be detected, suggesting that D3 has a protective role at is quite different from that in humans [117,118]. This could be due to these stages [110]. The number of cones, the photoreceptors for daylight the differences in the cellular distribution pattern of different TH trans- and colour vision, is greatly reduced in the retina of adult D3KO mice. porters between human and rodent brain [119,120] and shows the need

This has been linked to an excessive T3 signalling via TRβ2, leading to for additional animal models. Our knowledge on TH transporters in a high but transient increase in cell death of cones in the first postnatal non-mammalian vertebrates has started to grow during the last years V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141 137 and provides some interesting perspectives in order to fully understand In amphibians, the expression pattern of TH transporters has been theimpactofMCT8deficiency in developing human brain. studied by RT-PCR in different tissues of metamorphosing X. tropicalis from pre-metamorphosis towards the end of metamorphic climax (NF53 to NF64) [124].Ingeneral,mct8 and mct10 seem to be most abun- 4.1. Non-mammalian pre-juvenile stages dantly expressed but the temporal expression pattern of most transporters varies between tissues, in correlation with the timing of their The first, and to our knowledge, the only TH transporter that has metamorphic changes [124] (Fig. 6). Growth in hindlimbs seems to be been functionally characterized in fish is the zebrafish Mct8 [121].An associated with high levels of mct8 and mct10 and transient peaks in interesting feature is that although zebrafish Mct8 transports T4 and oatp1c1 and lat1 mRNA. The expression of mct8 in tail increases strongly T3 when tested in cell culture at 37 °C, it only transports T3 when tested throughout its regression and also mct10 and lat1 mRNA levels increase. at 26 °C, which is close to the normal body temperature in zebrafish. In gill mct8, mct10 and oatp1c1 expression only increase towards the last The authors showed that the mct8 gene is expressed in a wide variety stages of resorption. mct8, mct10 and lat1 expression increase in meta- of tissues in adult zebrafish and that mct8 mRNA is also present in ex- morphosing kidney, while metamorphosis in liver is accompanied by tracts of whole embryos, with a peak at 48 hpf [121]. Results of whole a strong decrease in mct8 mRNA. It is surprising that no major changes mount ISH show that mct8 mRNA is ubiquitously present in embryos have been observed in TH transporter expression in the metamorphos- at 10 hpf, but gradually becomes restricted to the brain and spinal ing brain, but they may occur in a region-specific way and therefore not cord by 48 hpf [23]. With the same technique, expression of oatp1c1 at be detectable in whole brain extracts [124]. It can be concluded that 48 hpf is only observed in vascular structures within the brain, while changes in cellular TH uptake and efflux are indeed another important mct10 expression is limited to the liver and the trigeminal ganglia level for the local regulation of nuclear TR-mediated signalling during [23]. Colocalisation studies of Mct8 and different cell-specific markers amphibian metamorphosis. It would certainly be interesting to start ex- in transgenic zebrafish larvae, carrying a fluorescent mct8 reporter con- periments with KD of transporters, especially Mct8 and Mct10, in this struct, indicate that mct8 is expressed in neurons and oligodendrocytes developmental model. but not in astrocytes, similar to the situation in mice. mct8 is also Chicken OATP1C1 has been cloned and characterized some years expressed in blood and lymph vessels in the trunk, in several brain ago, showing that it is a T4-specific transporter [125]. Our own group re- veins and blood vessels surrounding the brain, but not in the arteries cently cloned MCT8, MCT10, and LAT1 and functional characterization is at the level of the blood–brain barrier nor in the choroid plexus, which ongoing. Chickens, as well as some other investigated birds, seem to lack is different from the situation in rodents [23]. the gene for LAT2 (own unpublished results). We have investigated the The same research group has used morpholino KD to study the role distribution pattern of MCT8 and OATP1C1 in early chicken brain by RT- of Mct8 in zebrafish development. Their results show that at 48 hpf, PCR. Both transporters are expressed from the earliest stages studied Mct8 morphants have a deformed brain, spinal cord and notochord, (E4) but while MCT8 mRNA increases in all major brain regions from while the development of muscles and the vascular system seems unal- E4 up to E10, OATP1C1 mRNA in telencephalon and diencephalon tered [23]. In contrast, using the same approach, another research group shows the reverse pattern [65]. This agrees with our ISH results, show- recently found that next to defects in brain development, Mct8 KD also ing that MCT8 is widely expressed in neurons throughout the brain results in clear abnormalities in vascularisation, particularly at the level while clear expression of OATP1C1 is restricted to specific areas, such of the eye and the brain. In addition, their Mct8 morphants show a clear- as choroid plexus, hypothalamus and adenohypophysis. Unlike in ly reduced locomotor activity [122]. Also recently, the expression pat- zebrafish, MCT8 mRNA is also found in developing choroid plexus, but tern of several TH transporters has been reported in another teleost similar to zebrafish, it is absent from the microvessels in the brain fish, the fathead minnow (Pimephales promelas), but only at the adult forming the blood–brain barrier [66]. The only TH transporter present stage [123]. at the blood–brain barrier in embryonic chicken seems to be LAT1

Fig. 6. Ontogenetic proﬁle of mRNA levels for TH transporters (mct8, mct10, oatp1c1 and lat1) in different tissues of metamorphosing clawed frog (Xenopus tropicalis). Data are presented as fold change relative to the lowest level measured for each separate gene. The start of rapid hindlimb growth is accompanied by peaks in the expression of all transporters. Lat1 mRNA peaks early in gill resorption while the other transporters peak at the end. The expression of transporters in the resorbing tail shows a variable pattern. Developmental stages are indicated according to Nieuwkoop and Faber [28] (NF-stages 53 to 64) and representative images of metamorphic changes in hindlimb and tail have been taken from [28] (digitized on Xenbase) and [140] respectively. Data are taken from [124]. 138 V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141

(own unpublished results). In developing eye, MCT8 is widely expressed for LAT1. LAT2 is most abundant in microglia and also present in astro- in the retina (Fig. 5) and levels increase from E4 to E8. In de developing cytes and neurons, but not in oligodendrocytes and endothelial cells ear, the sites of MCT8 expression colocalize with those of DIO2, suggest- [130]. MCT8 and OATP1C1 proteins have been detected in the cerebral ing local cooperative actions between T3 production and transport. ISH cortex and at the level of the choroid plexus in rat foetuses at E21 did not reveal expression of OATP1C1 in developing eye or ear [66]. [131]. The functional implications of the specific cellular distribution pattern of each TH transporter in the developing rodent brain is only 4.2. Prenatal and neonatal mouse partly understood, but it can be assumed that they function both individ- ually and cooperatively in distributing active and inactive iodothyronines Mct8, Lat1,andLat2, but not Oatp1c1 mRNA have been detected by amongst proliferating and maturing brain cell populations. qPCR in femurs of prenatal mice from the first stage studied, namely The cell-specific expression pattern of the different TH transporters 14.5-day-old embryos (E14.5). Mct8 expression gradually decreases as in developing mouse cochlea suggests that, together with the Ds, they bone development progresses while expression of the other two trans- are important in timing the access of T3 to the differentiating cells. porters remains stable, a pattern suggesting that MCT8 may be an im- LAT1 is strongly expressed in cochlear blood vessels and may facilitate portant regulator of bone development and osteoblast differentiation uptake of THs from the general circulation. OATP1C1 colocalizes with

[126]. MCT8 is also expressed in mouse placenta and MCT8 deficiency D2 in fibrocytes and may stimulate uptake of T4 for conversion into T3. provokes some subtle changes in foetoplacental growth [127]. Studies MCT8 expression partly overlaps with TRβ, the predominant TR in the in juvenile/adult mice have also shown interesting expression patterns cochlea, and may be needed for cellular uptake of T3 and subsequent for TH transporters in other peripheral tissues such as kidney and intes- liganded TR-mediated action [133]. Data on TH transporter expression tine [128], but data from prenatal and early postnatal stages are lacking. in the mouse eye are still missing, but given the recent data on MCT8 ex- A dynamic expression pattern for TH transporters is present in de- pression in embryonic chicken retina (see above), they will almost cer- veloping mouse brain. MCT8 was first studied by ISH, showing predom- tainly have a regulatory role in the development of that sensory system inantly neuronal Mct8 expression in the forebrain at E18 [129].AqPCR too. study in developing mouse brain from E15 up to postnatal day 20 KO mice have been created for all known TH transporters. So far they (P20) showed that mRNA levels for Mct8, Oatp1c1, Lat1 and Lat2 in hip- have been studied predominantly in juvenile/adult stages. Interestingly, pocampus increase during embryogenesis, continue to rise until P5 and a recent study in perinatal MCT8KO mice reveals some striking differ- then decline again towards P20, while expression of Mct10 remains ences with the adult phenotype. At the peripheral level, adult stable (Fig. 7). Similarly, a peak is present in Mct8, Lat1 and Lat2 expres- MCT8KO mice have reduced plasma T4 but increased plasma T3 and sion in neocortex and cerebellum around days P5–P10 [130]. For com- liver T3 content. In brain, T4 and T3 content are decreased, accompanied parison, qPCR on rat cerebral cortex from late foetal to early postnatal by an increase in D2 and a decrease in D3 as a compensatory response to stages also shows a postnatal decrease in Mct8, but an increase in the hypothyroid status [117,118]. Surprisingly, perinatal MCT8KO mice

Oatp1c1 and Lat2 expression [131]. Results from a transgenic mouse have higher plasma T4 than wild type littermates from E18 up to P0 and line carrying an Oatp1c1 reporter construct indicate that Oatp1c1 is tran- thereafter plasma T4 levels in KO and wild type pups are similar at least siently expressed in neuronal progenitors in embryonic mouse cortex up to P7. This transient increase is accompanied by higher tissue content but absent in adult neurons, while its expression in blood vessels and of T4 and T3 and higher expression levels of some TH-responsive genes choroid plexus is sustained from embryonic development to adulthood in both cerebral cortex and liver. The fact that this hyperthyroid status in [132]. The cell-speciﬁc distribution of LAT1, LAT2 and MCT8 has been brain is accompanied by increased expression of Oatp1c1, Lat1 and/or studied at the protein level in primary cultures of different cell types Lat2 suggests that these transporters compensate for the lack of MCT8 from E16 mouse brain. MCT8 is most abundant in neurons, but detect- and increase TH uptake into the brain [134]. Indeed, the recent descrip- able in all cell types except microglia. The distribution pattern is similar tion of a MCT8/OATP1C1 double KO mouse showed that these mice suf- fer from impaired and delayed brain development, indicative of a severe hypothyroid status [135]. The above results illustrate that due to the interaction of ontogenetic changes at different levels of the thyroid system, TH availability and action may be substantially different in developing compared to mature KO animals, as has also been found before in developing D3KO mouse [96].

5. Conclusion

The tissue- and stage-speciﬁc expression patterns of Ds observed in the development of different vertebrates are a strong indication that TH activation and inactivation by these enzymes is an essential step in con-

trolling the intracellular availability of T3 (and in teleosts also 3,5-T2) and hence TR-mediated signalling. Studies following D knockout/knockdown or overexpression in three different vertebrate models, namely zebrafish, Xenopus and mouse, have now also provided functional proof for the involvement of Ds in the development of a variety of organs. It is interesting to notice that in general, deficiency of the TH-inactivating enzyme D3 seems to result in the most severe phenotype, suggesting that a premature switch from unliganded to liganded TR action is often Fig. 7. Ontogenetic profile of mRNA levels for TH transporters (mct8, oatp1c1, lat1 and lat2) in mouse hippocampus from embryonic day 15 (E15) up to postnatal day 20 (P20). Data more detrimental than a delayed one. In this regard it would certainly are presented as fold change relative to the lowest level measured for each separate gene. be interesting to study in more detail early embryonic stages in KO mct8 mRNA levels peak shortly before birth while oatp1c1 and lat2 expression continue to mice, since studies in other vertebrate models are underlining the impact rise up to one week after birth. mRNA levels for mct10 remain stable throughout this pe- fi of changes in these early stages on further development and later life. It is riod and are not included in the gure. fi Representative images for the developmental changes occurring at the level of the hippo- also interesting to see that many cell-speci c expression patterns have campus have been taken from the Allen Brain Atlas portal (Allen Institute for Brain Science) been conserved throughout vertebrate evolution, as found for instance [141]. Data are taken from [130]. for eye and ear development where changes in Ds seem to be linked to V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141 139 changes in TRβ action. Our knowledge on the distribution and function of [21] W. Dong, et al., Using whole mount in situ hybridization to examine thyroid hormone deiodinase expression in embryonic and larval zebrafish: a tool for examin- TH transporters is also increasing rapidly. As a consequence of the link ing OH-BDE toxicity to early life stages, Aquat. Toxicol. 132–133 (2013) 190–199. found in humans between MCT8 mutation and the severe AHD syn- [22] C. Thisse, et al., Spatial and temporal expression patterns of selenoprotein genes drome, research on the role of TH transporters so far mainly focused on during embryogenesis in zebrafish, Gene Expr. Patterns 3 (4) (2003) 525–532. [23] G.D. Vatine, et al., Zebrafish as a model for monocarboxyl transporter 8-deficiency, brain development. We should however not neglect their role in devel- J. Biol. Chem. 288 (1) (2013) 169–180. opment of other structures and more research, including studies in meta- [24] V.M. Bedell, S.E. Westcot, S.C. Ekker, Lessons from morpholino-based screening in morphosing species, is needed. zebrafish, Brief. Funct Genom. 10 (4) (2011) 181–188. Although substantial information on the ontogenetic distribution [25] C.N. Walpita, et al., Type 2 iodothyronine deiodinase is essential for thyroid hormone-dependent embryonic development and pigmentation in zebrafish, pattern and function of TH receptors, deiodinases, and TH transporters Endocrinology 150 (1) (2009) 530–539. is now available, we are far from exactly understanding how they [26] C.N. Walpita, A.D. Crawford, V.M. Darras, Combined antisense knockdown of type 1 work together in regulating TR-dependent gene repression and activa- and type 2 iodothyronine deiodinases disrupts embryonic development in zebrafish (Danio rerio), Gen. Comp. Endocrinol. 166 (1) (2010) 134–141. tion in developing tissues. Studies in vertebrates where whole body ex- [27] B.L. Bohnsack, A. Kahana, Thyroid hormone and retinoic acid interact to regulate pression of one or more of the players has been silenced by gene zebrafish craniofacial neural crest development, Dev. Biol. 373 (2) (2013) 300–309. knockout/knockdown have started to provide essential pieces of the [28] P.D. Nieuwkoop, J. Faber, Normal Table of Xenopus laevis (Daudin): A Systematical and Chronological Survey of the Development from the Fertilized Egg Till the End puzzle. The next and essential step is to create models with conditional of Metamorphosis, Garland Pub., New York, 1994. 10 (252 pp., leaves of plates). silencing that can be restricted in a tissue- and time-specific way. This is [29] G. Morvan Dubois, et al., Deiodinase activity is present in Xenopus laevis during already ongoing in mice and with the recent progress in molecular tech- early embryogenesis, Endocrinology 147 (10) (2006) 4941–4949. [30] J.B. Fini, et al., Thyroid hormone signaling in the Xenopus laevis embryo is func- niques other vertebrate models will hopefully follow soon. tional and susceptible to endocrine disruption, Endocrinology 153 (10) (2012) 5068–5081. [31] A.J. Tindall, et al., Expression of enzymes involved in thyroid hormone metabolism References during the early development of Xenopus tropicalis, Biol. Cell. 99 (3) (2007) 151–163. [32] G. Morvan-Dubois, B.A. Demeneix, L.M. Sachs, Xenopus laevis as a model for study- [1] F. Flamant, K. Gauthier, Thyroid hormone receptors: the challenge of elucidating ing thyroid hormone signalling: from development to metamorphosis, Mol. Cell. isotype-specificfunctionsandcell-specific response, Biochim. Biophys. Acta 1830 Endocrinol. 293 (1–2) (2008) 71–79. (7) (2013) 3900–3907. [33] D.E. Banker, J. Bigler, R.N. Eisenman, The thyroid hormone receptor gene (c-erbA [2] S.Y. Cheng, J.L. Leonard, P.J. Davis, Molecular aspects of thyroid hormone actions, alpha) is expressed in advance of thyroid gland maturation during the early em- Endocr. Rev. 31 (2) (2010) 139–170. bryonic development of Xenopus laevis, Mol. Cell. Biol. 11 (10) (1991) 5079–5089. [3] A. Mendoza, et al., 3,5-T2 is an alternative ligand for the thyroid hormone receptor [34] E. Havis, et al., Unliganded thyroid hormone receptor is essential for Xenopus laevis beta1, Endocrinology 154 (8) (2013) 2948–2958. eyedevelopment,EMBOJ.25(20)(2006)4943–4951. [4] A. Orozco, et al., 3,5-Diiodothyronine (T2) is on a role. A new hormone in search of [35] G. Morvan-Dubois, J.B. Fini, B.A. Demeneix, Is thyroid hormone signaling relevant recognition, Gen. Comp. Endocrinol. (2014), http://dx.doi.org/10.1016/j.ygcen. for vertebrate embryogenesis? Curr. Top. Dev. Biol. 103 (2013) 365–396. 2014.02.14 [Epub ahead of print]. [36] A. Yin, et al., Wnt signaling is required for early development of zebrafish [5] M. Harada, et al., cDNA cloning and expression analysis of thyroid hormone recep- swimbladder, PLoS One 6 (3) (2011) e18431. tor in the coho salmon Oncorhynchus kisutch during smoltification, Gen. Comp. [37] C.L. Winata, et al., Development of zebrafish swimbladder: the requirement of Endocrinol.155(3)(2008)658–667. hedgehog signaling in specification and organization of the three tissue layers, [6] L. Fairclough, J.R. Tata, An immunocytochemical analysis of the expression of Dev. Biol. 331 (2) (2009) 222–236. thyroid hormone receptor alpha and beta proteins during natural and thyroid [38] L. Wang, Y.Y. Shao, R.T. Ballock, Thyroid hormone interacts with the Wnt/beta-catenin hormone-induced metamorphosis in Xenopus, Develop. Growth Differ. 39 (3) signaling pathway in the terminal differentiation of growth plate chondrocytes, (1997) 273–283. J. Bone Miner. Res. 22 (12) (2007) 1988–1995. [7] D. Forrest, M. Sjoberg, B. Vennstrom, Contrasting developmental and tissue-specific [39] A. Ishizuya-Oka, et al., Thyroid hormone-induced expression of sonic hedgehog expression of alpha and beta thyroid hormone receptor genes, EMBO J. 9 (5) (1990) correlates with adult epithelial development during remodeling of the Xenopus 1519–1528. stomach and intestine, Differentiation 69 (1) (2001) 27–37. [8] F. Flamant, J. Samarut, Thyroid hormone receptors: lessons from knockout and [40] M. Dentice, Hedgehog-mediated regulation of thyroid hormone action through knock-in mutant mice, Trends Endocrinol. Metab. 14 (2) (2003) 85–90. iodothyronine deiodinases, Expert Opin. Ther. Targets 15 (4) (2011) 493–504. [9] P.M. Yen, et al., Effects of ligand and thyroid hormone receptor isoforms on hepatic [41] Y.W. Liu, W.K. Chan, Thyroid hormones are important for embryonic to larval tran- gene expression profiles of thyroid hormone receptor knockout mice, EMBO Rep. 4 sitory phase in zebrafish, Differentiation 70 (1) (2002) 36–45. (6) (2003) 581–587. [42] V.A. Galton, S.H. Ingbar, Observations on the relation between the action and the [10] A.C. Bianco, et al., Biochemistry, cellular and molecular biology, and physio- degradation of thyroid hormones as indicated by studies in the tadpole and the logical roles of the iodothyronine selenodeiodinases, Endocr. Rev. 23 (1) frog, Endocrinology 70 (1962) 622–632. (2002) 38–89. [43] V.A. Galton, The role of thyroid-hormone in amphibian development, Am. Zool. 28 [11] A. Orozco, et al., Iodothyronine deiodinases: a functional and evolutionary perspec- (2)(1988)309–318. tive, J. Endocrinol. 215 (2) (2012) 207–219. [44] Z. Wang, D.D. Brown, Thyroid hormone-induced gene expression program for am- [12] V.M. Darras, S.L. Van Herck, Iodothyronine deiodinase structure and function: from phibian tail resorption, J. Biol. Chem. 268 (22) (1993) 16270–16278. ascidians to humans, J. Endocrinol. 215 (2) (2012) 189–206. [45] D.L. St Germain, et al., A thyroid hormone-regulated gene in Xenopus laevis encodes [13] A.L. Maia, et al., Deiodinases: the balance of thyroid hormone: type 1 iodothyronine a type III iodothyronine 5-deiodinase, Proc. Natl. Acad. Sci. U. S. A. 91 (16) (1994) deiodinase in human physiology and disease, J. Endocrinol. 209 (3) (2011) 7767–7771. 283–297. [46] K.B. Becker, et al., The type 2 and type 3 iodothyronine deiodinases play important [14] M.J. Schneider, et al., Targeted disruption of the type 1 selenodeiodinase gene (Dio1) roles in coordinating development in Rana catesbeiana tadpoles, Endocrinology results in marked changes in thyroid hormone economy in mice, Endocrinology 147 138 (7) (1997) 2989–2997. (1) (2006) 580–589. [47] L. Cai, D.D. Brown, Expression of type II iodothyronine deiodinase marks the time [15] O. Elsalini, Zebrafish hhex, nk2.1a, and pax2.1 regulate thyroid growth and differ- that a tissue responds to thyroid hormone-induced metamorphosis in Xenopus entiation downstream of Nodal-dependent transcription factors, Dev. Biol. 263 (1) laevis, Dev. Biol. 266 (1) (2004) 87–95. (2003) 67–80. [48] D.L. Berry, et al., The expression pattern of thyroid hormone response genes in re- [16] K. Itoh, et al., Three members of the iodothyronine deiodinase family, dio1, modeling tadpole tissues defines distinct growth and resorption gene expression dio2 and dio3, are expressed in spatially and temporally specific patterns dur- programs, Dev. Biol. 203 (1) (1998) 24–35. ing metamorphosis of the flounder, Paralichthys olivaceus, Zool. Sci. 27 (7) [49] D.D. Brown, The role of deiodinases in amphibian metamorphosis, Thyroid 15 (8) (2010) 574–580. (2005) 815–821. [17] M. Heijlen, et al., Knockdown of type 3 iodothyronine deiodinase severely perturbs [50] H. Huang, N. Marsh-Armstrong, D.D. Brown, Metamorphosis is inhibited in trans- both embryonic and early larval development in zebrafish, Endocrinology 155 (4) genic Xenopus laevis tadpoles that overexpress type III deiodinase, Proc. Natl. (2014) 1547–1559. Acad. Sci. U. S. A. 96 (3) (1999) 962–967. [18] V.M. Darras, et al., Thyroid hormone receptors in two model species for vertebrate [51] N. Marsh-Armstrong, et al., Asymmetric growth and development of the Xenopus embryonic development: chicken and zebrafish, J. Thyroid. Res. 2011 (2011) laevis retina during metamorphosis is controlled by type III deiodinase, Neuron 402320. 24 (4) (1999) 871–878. [19] C.N. Walpita, et al., The effect of 3,5,3′-triiodothyronine supplementation on [52] O. Marchand, et al., Molecular cloning and developmental expression patterns of zebrafish (Danio rerio) embryonic development and expression of iodothyronine thyroid hormone receptors and T3 target genes in the turbot (Scophtalmus deiodinases and thyroid hormone receptors, Gen. Comp. Endocrinol. 152 (2–3) maximus) during post-embryonic development, Gen. Comp. Endocrinol. 135 (3) (2007) 206–214. (2004) 345–357. [20] M. Heijlen, A.M. Houbrechts, V.M. Darras, Zebrafish as a model to study peripheral [53] P.H. Klaren, et al., The thyroid gland and thyroid hormones in Senegalese sole thyroid hormone metabolism in vertebrate development, Gen. Comp. Endocrinol. (Solea senegalensis) during early development and metamorphosis, Gen. Comp. 188 (2013) 289–296. Endocrinol. 155 (3) (2008) 686–694. 140 V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141

[54] E. Isorna, et al., Iodothyronine deiodinases and thyroid hormone receptors regula- [85] J.H. Dussault, F. Labrie, Development of the hypothalamic–pituitary–thyroid axis in tion during flatfish (Solea senegalensis) metamorphosis, J. Exp. Zool. B Mol. Dev. the neonatal rat, Endocrinology 97 (5) (1975) 1321–1324. Evol. 312B (3) (2009) 231–246. [86] D.A. Fisher, et al., Ontogenesis of hypothalamic–pituitary–thyroid function and [55] A.M. Schreiber, J.L. Specker, Metamorphosis in the summer flounder (Paralichthys metabolism in man, sheep, and rat, Recent Prog. Horm. Res. 33 (1976) 59–116. dentatus): stage-specific developmental response to altered thyroid status, Gen. [87] A. Hernandez, et al., The gene locus encoding iodothyronine deiodinase type 3 Comp. Endocrinol. 111 (2) (1998) 156–166. (Dio3) is imprinted in the fetus and expresses antisense transcripts, Endocrinology [56] M.A. Campinho, et al., Coordination of deiodinase and thyroid hormone receptor 143 (11) (2002) 4483–4486. expression during the larval to juvenile transition in sea bream (Sparus aurata, [88] W. Reik, J. Walter, Genomic imprinting: parental influence on the genome, Nat. Linnaeus), Gen. Comp. Endocrinol. 165 (2) (2010) 181–194. Rev. Genet. 2 (1) (2001) 21–32. [57] F.M. McNabb, Avian thyroid development and adaptive plasticity, Gen. Comp. [89] M. Charalambous, A. Hernandez, Genomic imprinting of the type 3 thyroid hor- Endocrinol. 147 (2) (2006) 93–101. mone deiodinase gene: regulation and developmental implications, Biochim. [58] F.M.A. Mcnabb, Comparative thyroid development in precocial Japanese-quail and Biophys. Acta 1830 (7) (2013) 3946–3955. altricial ring doves, J. Exp. Zool. (1987) 281–290. [90] E. Roti, et al., Human placenta is an active site of thyroxine and 3,3′5-triiodothyro- [59] R.C. Thommes, et al., Immunocytochemical demonstration of T4 content and TSH- nine tyrosyl ring deiodination, J. Clin. Endocrinol. Metab. 53 (3) (1981) 498–501. binding by cells of the thyroid of the developing chick embryo, Gen. Comp. [91] G. Morreale de Escobar, et al., Transfer of thyroxine from the mother to the rat Endocrinol. 85 (1) (1992) 79–85. fetus near term: effects on brain 3,5,3′-triiodothyronine deficiency, Endocrinology [60] M. Borges, J. LaBourene, S.H. Ingbar, Changes in hepatic iodothyronine metabolism 122 (4) (1988) 1521–1531. during ontogeny of the chick embryo, Endocrinology 107 (6) (1980) 1751–1761. [92] V.A. Galton, The roles of the iodothyronine deiodinases in mammalian develop- [61] V.A. Galton, A. Hiebert, The ontogeny of the enzyme systems for the 5′- and 5- ment, Thyroid 15 (8) (2005) 823–834. deiodination of thyroid hormones in chick embryo liver, Endocrinology 120 (6) [93] J.M. Bates, D.L. St Germain, V.A. Galton, Expression profiles of the three (1987) 2604–2610. iodothyronine deiodinases, D1, D2, and D3, in the developing rat, Endocrinology [62] Y. Dickstein, et al., The metabolism of T4 and T3 in cultured chick-embryo heart 140 (2) (1999) 844–851. cells, Mol. Cell. Endocrinol. 20 (1) (1980) 45–57. [94] V.A. Galton, et al., Life without thyroxine to 3,5,3′-triiodothyronine conversion: [63] V.M. Darras, et al., Ontogeny of type I and type III deiodinase activities in embryon- studies in mice devoid of the 5′-deiodinases, Endocrinology 150 (6) (2009) ic and posthatch chicks: relationship with changes in plasma triiodothyronine and 2957–2963. growth hormone levels, Comp. Biochem. Physiol. Comp. Physiol. 103 (1) (1992) [95] M.J. Schneider, et al., Targeted disruption of the type 2 selenodeiodinase gene 131–136. (DIO2) results in a phenotype of pituitary resistance to T4, Mol. Endocrinol. 15 [64] S. Van der Geyten, et al., Differential expression of iodothyronine deiodinases in (12) (2001) 2137–2148. chicken tissues during the last week of embryonic development, Gen. Comp. [96] A. Hernandez, et al., Type 3 deiodinase is critical for the maturation and function of Endocrinol. 128 (1) (2002) 65–73. the thyroid axis, J. Clin. Invest. 116 (2) (2006) 476–484. [65] S.L. Van Herck, et al., Expression profile and thyroid hormone responsiveness of [97] J.A. Hall, et al., Absence of thyroid hormone activation during development under- transporters and deiodinases in early embryonic chicken brain development, lies a permanent defect in adaptive thermogenesis, Endocrinology 151 (9) (2010) Mol. Cell. Endocrinol. 349 (2) (2012) 289–297. 4573–4582. [66] S. Geysens, et al., Dynamic mRNA distribution pattern of thyroid hormone [98] M. Dentice, et al., The FoxO3/type 2 deiodinase pathway is required for normal transporters and deiodinases during early embryonic chicken brain development, mouse myogenesis and muscle regeneration, J. Clin. Invest. 120 (11) (2010) Neuroscience 221 (2012) 69–85. 4021–4030. [67] B. Gereben, et al., Ontogenic redistribution of type 2 deiodinase messenger ribonu- [99] R. Ambrosio, et al., Epigenetic control of type 2 and 3 deiodinases in myogenesis: cleic acid in the brain of chicken, Endocrinology 145 (8) (2004) 3619–3625. role of lysine-specific demethylase enzyme and FoxO3, Nucleic Acids Res. 41 (6) [68] C.H. Verhoelst, S.A. Roelens, V.M. Darras, Role of spatiotemporal expression of (2013) 3551–3562. iodothyronine deiodinase proteins in cerebellar cell organization, Brain Res. Bull. [100] J.H. Bassett, et al., Optimal bone strength and mineralization requires the type 2 67(3)(2005)196–202. iodothyronine deiodinase in osteoblasts, Proc. Natl. Acad. Sci. U. S. A. 107 (16) [69] C.H. Verhoelst, et al., Type II iodothyronine deiodinase protein in chicken choroid (2010) 7604–7609. plexus: additional perspectives on T3 supply in the avian brain, J. Endocrinol. [101] A.J. Williams, et al., Iodothyronine deiodinase enzyme activities in bone, Bone 43 183 (1) (2004) 235–241. (1) (2008) 126–134. [70] D. Forrest, et al., Distinct functions for thyroid hormone receptors alpha and beta in [102] L.P. Capelo, et al., Deiodinase-mediated thyroid hormone inactivation minimizes brain development indicated by differential expression of receptor genes, EMBO J. thyroid hormone signaling in the early development of fetal skeleton, Bone 43 10 (2) (1991) 269–275. (5) (2008) 921–930. [71] J.M. Trimarchi, et al., Thyroid hormone components are expressed in three [103] M.C. Medina, et al., The thyroid hormone-inactivating type III deiodinase is sequential waves during development of the chick retina, BMC Dev. Biol. 8 expressed in mouse and human beta-cells and its targeted inactivation impairs (2008) 101. insulin secretion, Endocrinology 152 (10) (2011) 3717–3727. [72] A.H. Conrad, et al., Thyroxine affects expression of KSPG-related genes, the carbon- [104] A. Marsili, et al., Mice with a targeted deletion of the type 2 deiodinase are insulin ic anhydrase II gene, and KS sulfation in the embryonic chicken cornea, Invest. resistant and susceptible to diet induced obesity, PLoS One 6 (6) (2011) e20832. Ophthalmol. Vis. Sci. 47 (1) (2006) 120–132. [105] V.A. Galton, et al., Thyroid hormone homeostasis and action in the type 2 [73] C.H. Verhoelst, S. Van Der Geyten, V.M. Darras, Renal and hepatic distribution of deiodinase-deficient rodent brain during development, Endocrinology 148 (7) type I and type III iodothyronine deiodinase protein in chicken, J. Endocrinol. 181 (2007) 3080–3088. (1) (2004) 85–90. [106] R.P. Peeters, et al., Cerebellar abnormalities in mice lacking type 3 deiodinase [74] M. Dentice, et al., The hedgehog-inducible ubiquitin ligase subunit WSB-1 modu- and partial reversal of phenotype by deletion of thyroid hormone receptor alpha1, lates thyroid hormone activation and PTHrP secretion in the developing growth Endocrinology 154 (1) (2013) 550–561.

plate, Nat. Cell Biol. 7 (7) (2005) 698–705. [107] L. Ng, M.W. Kelley, D. Forrest, Making sense with thyroid hormone—the role of T3 [75] S. Van der Geyten, et al., Transcriptional regulation of iodothyronine deiodinases in auditory development, Nat. Rev. Endocrinol. 9 (5) (2013) 296–307. during embryonic development, Mol. Cell. Endocrinol. 183 (1–2) (2001) 1–9. [108] L. Ng, et al., Hearing loss and retarded cochlear development in mice lacking type 2 [76] V.M. Darras, et al., Plasma thyroid hormone levels and iodothyronine deiodinase iodothyronine deiodinase, Proc. Natl. Acad. Sci. U. S. A. 101 (10) (2004) activity following an acute glucocorticoid challenge in embryonic compared with 3474–3479. posthatch chickens, Gen. Comp. Endocrinol. 104 (2) (1996) 203–212. [109] L. Ng, et al., A protective role for type 3 deiodinase, a thyroid hormone-inactivating [77] V.M. Darras, et al., Growth hormone acutely decreases type III iodothyronine enzyme, in cochlear development and auditory function, Endocrinology 150 (4) deiodinase in chicken liver, FEBS Lett. 310 (1) (1992) 5–8. (2009) 1952–1960. [78] Y. Tanabe, N. Saito, T. Nakamura, Ontogenetic steroidogenesis by testes, ovary, and [110] L. Ng, et al., Type 3 deiodinase, a thyroid-hormone-inactivating enzyme, controls adrenals of embryonic and postembryonic chickens (Gallus domesticus), Gen. survival and maturation of cone photoreceptors, J. Neurosci. 30 (9) (2010) Comp. Endocrinol. 63 (3) (1986) 456–463. 3347–3357. [79] G.E. Reyns, et al., Regulation of thyroid hormone availability in liver and brain by [111] A. Hernandez, et al., Type 3 deiodinase deﬁciency causes spatial and temporal al- glucocorticoids, Gen. Comp. Endocrinol. 140 (2) (2005) 101–108. terations in brain T3 signaling that are dissociated from serum thyroid hormone [80] S. Van der Geyten, V.M. Darras, Developmentally deﬁned regulation of thyroid levels, Endocrinology 151 (11) (2010) 5550–5558. hormone metabolism by glucocorticoids in the rat, J. Endocrinol. 185 (2) (2005) [112] A. Hernandez, et al., Critical role of types 2 and 3 deiodinases in the negative reg-

327–336. ulation of gene expression by T3 in the mouse cerebral cortex, Endocrinology 153 [81] V.M. Darras, et al., Effects of dexamethasone treatment on iodothyronine (6)(2012)2919–2928. deiodinase activities and on metamorphosis-related morphological changes in [113] E.C. Friesema, et al., Thyroid hormone transporters, Vitam. Horm. 70 (2005) the axolotl (Ambystoma mexicanum), Gen. Comp. Endocrinol. 127 (2) (2002) 137–167. 157–164. [114] W.E. Visser, et al., Thyroid hormone transport in and out of cells, Trends [82] J.S. LoPresti, et al., Alterations in 3,3′5′-triiodothyronine metabolism in response to Endocrinol. Metab. 19 (2) (2008) 50–56. propylthiouracil, dexamethasone, and thyroxine administration in man, J. Clin. [115] E.C. Friesema, et al., Association between mutations in a thyroid hormone trans- Invest. 84 (5) (1989) 1650–1656. porter and severe X-linked psychomotor retardation, Lancet 364 (9443) (2004) [83] S. Harpavat, C.L. Cepko, RCAS-RNAi: a loss-of-function method for the developing 1435–1437. chick retina, BMC Dev. Biol. 6 (2006) 2. [116] A.M. Dumitrescu, et al., A novel syndrome combining thyroid and neurological ab- [84] J. Gross, C.P. Leblond, Metabolism of the thyroid hormone in the rat as shown by normalities is associated with mutations in a monocarboxylate transporter gene, physiological doses of labeled thyroxine, J. Biol. Chem. 184 (2) (1950) 489–500. Am. J. Hum. Genet. 74 (1) (2004) 168–175. V.M. Darras et al. / Biochimica et Biophysica Acta 1849 (2015) 130–141 141

[117] A.M. Dumitrescu, et al., Tissue-specific thyroid hormone deprivation and excess [129] H. Heuer, et al., The monocarboxylate transporter 8 linked to human psychomotor in monocarboxylate transporter (mct) 8-deficient mice, Endocrinology 147 (9) retardation is highly expressed in thyroid hormone-sensitive neuron populations, (2006) 4036–4043. Endocrinology 146 (4) (2005) 1701–1706. [118] M. Trajkovic, et al., Abnormal thyroid hormone metabolism in mice lacking the [130] D. Braun, et al., Developmental and cell type-specific expression of thyroid hor- monocarboxylate transporter 8, J. Clin. Invest. 117 (3) (2007) 627–635. mone transporters in the mouse brain and in primary brain cells, Glia 59 (3) [119] L.M. Roberts, et al., Expression of the thyroid hormone transporters monocarboxyl- (2011) 463–471. ate transporter-8 (SLC16A2) and organic ion transporter-14 (SLCO1C1) at the [131] C. Grijota-Martinez, et al., Lack of action of exogenously administered T3 on the blood–brain barrier, Endocrinology 149 (12) (2008) 6251–6261. fetal rat brain despite expression of the monocarboxylate transporter 8, Endocri- [120] E.K. Wirth, et al., Neuronal 3′,3,5-triiodothyronine (T3) uptake and behavioral phe- nology 152 (4) (2011) 1713–1721. notype of mice deficient in Mct8, the neuronal T3 transporter mutated in Allan– [132] M.F. Lang, et al., A transgenic approach to identify thyroxine transporter- Herndon–Dudley syndrome, J. Neurosci. 29 (30) (2009) 9439–9449. expressing structures in brain development, J. Endocrinol. 23 (12) (2011) [121] F.J. Arjona, et al., Identification and functional characterization of zebrafish solute 1194–1203. carrier Slc16a2 (Mct8) as a thyroid hormone membrane transporter, Endocrinolo- [133] D.S. Sharlin, T.J. Visser, D. Forrest, Developmental and cell-specific expression of gy 152 (12) (2011) 5065–5073. thyroid hormone transporters in the mouse cochlea, Endocrinology 152 (12) [122] E. de Vrieze, et al., Knockdown of monocarboxylate transporter 8 (mct8) disturbs (2011) 5053–5064. brain development and locomotion in zebrafish, Endocrinology (2014) en20131962. [134] A.M. Ferrara, et al., Changes in thyroid status during perinatal development of [123] A.M. Muzzio, et al., Tissue distribution and thyroid hormone effects on mRNA MCT8-deficient male mice, Endocrinology 154 (7) (2013) 2533–2541. abundance for membrane transporters Mct8, Mct10, and organic anion- [135] S. Mayerl, et al., Transporters MCT8 and OATP1C1 maintain murine brain thyroid transporting polypeptides (Oatps) in a teleost fish, Comp. Biochem. Physiol. A hormone homeostasis, J. Clin. Invest. (2014), http://dx.doi.org/10.1172/JCI70324 Mol. Integr. Physiol. 167 (2014) 77–89. [Epub ahead of print]. [124] K.A. Connors, et al., Characterization of thyroid hormone transporter expression [136] M. Heijlen, et al., Knockdown of type 3 iodothyronine deiodinase severely perturbs during tissue-specific metamorphic events in Xenopus tropicalis,Gen.Comp. both embryonic and early larval development in zebrafish, Endocrinology (2014) Endocrinol. 168 (1) (2010) 149–159. en20131660. [125] N. Nakao, et al., Possible involvement of organic anion transporting polypeptide [137] A.C. Taylor, J.J. Kollros, Stages in the normal development of Rana pipiens larvae, 1c1 in the photoperiodic response of gonads in birds, Endocrinology 147 (3) Anat. Rec. 94 (1946) 7–13. (2006) 1067–1073. [138] R. Rugh, The Frog; Its Reproduction and Development, Blakiston, Philadelphia, [126] L.P. Capelo, et al., The monocarboxylate transporter 8 and L-type amino acid trans- 1952. (x, 336 pp.). porters 1 and 2 are expressed in mouse skeletons and in osteoblastic MC3T3-E1 [139] M. Aritaki, T. Seikai, Temperature effects on early development and occurrence of cells, Thyroid 19 (2) (2009) 171–180. metamorphosis-related morphological abnormalities in hatchery-reared brown [127] E. Vasilopoulou, et al., Monocarboxylate transporter 8 modulates the viability and sole Pseudopleuronectes herzensteini, Aquaculture 240 (1–4) (2004) 517–530. invasive capacity of human placental cells and fetoplacental growth in mice, PLoS [140] M.H.I. Dodd, J.M. Dodd, The biology of metamorphosis, in: B. Lofts (Ed.), Physiology One 8 (6) (2013) e65402. of the Amphibia, Academic Press, New York, 1976. [128] D. Braun, et al., Aminoaciduria, but normal thyroid hormone levels and signalling, [141] A.M. Henry, J.G. Hohmann, High-resolution gene expression atlases for adult and in mice lacking the amino acid and thyroid hormone transporter Slc7a8, Biochem. developing mouse brain and spinal cord, Mamm. Genome 23 (9–10) (2012) J. 439 (2) (2011) 249–255. 539–549.