Tian et al. Nutrition & Metabolism (2018) 15:7 DOI 10.1186/s12986-017-0235-x RESEARCH Open Access The interaction of selenoprotein F (SELENOF) with retinol dehydrogenase 11 (RDH11) implied a role of SELENOF in vitamin A metabolism Jing Tian1* , Jiapan Liu1, Jieqiong Li2, Jingxin Zheng3, Lifang Chen4, Yujuan Wang1, Qiong Liu1 and Jiazuan Ni1 Abstract Background: Selenoprotein F (SELENOF, was named as 15-kDa selenoprotein) has been reported to play important roles in oxidative stress, endoplasmic reticulum (ER) stress and carcinogenesis. However, the biological function of SELENOF is still unclear. Methods: A yeast two-hybrid system was used to screen the interactive protein of SELENOF in a human fetal brain cDNA library. The interaction between SELENOF and interactive protein was validated by fluorescence resonance energy transfer (FRET), co-immunoprecipitation (co-IP) and pull-down assays. The production of retinol was detected by high performance liquid chromatograph (HPLC). Results: Retinol dehydrogenase 11 (RDH11) was found to interact with SELENOF. RDH11 is an enzyme for the reduction of all-trans-retinaldehyde to all-trans-retinol (vitamin A). The production of retinol was decreased by SELENOF overexpression, resulting in more retinaldehyde. Conclusions: SELENOF interacts with RDH11 and blocks its enzyme activity to reduce all-trans-retinaldehyde. Keywords: SELENOF (Seleonoprotein F) , Yeast two hybrid system, Protein-protein interaction, Retinol dehydrogenase 11 (RDH11), Fluorescence resonance energy transfer (FRET), Co-immunoprecipitation (co-IP), Pull- down, Retinol (vitamin a), Retinaldehyde Background SELENOF shows that the protein contains a Selenium (Se) is a necessary trace element for human thioredoxin-like motif. The redox potential of this motif health. It primarily exerts its function through seleno- is between that of thioredoxin and protein disulfideisome- proteins in which Se is present in the form of selenocys- rase, so SELENOF was assumed to be a thiol-disulfide teine (Sec), which is encoded by UGA, traditionally read oxido-reductase and/or isomerase [3]. The N-terminal as a stop codon in the coding region. SELENOF, initially signal peptide of SELENOF guides it to localize to the named as 15-kDa selenoprotein (Sep15) is one of the 25 ER [4]. Mouse SELENOF was shown to form a 1:1 genes in humans encoding for selenoproteins [1]. It is complex with UDP-glucose: glycoprotein glucosyl- conserved from worms to human beings, whereas the transferase 1 (UGGT1), which is a chaperone involved genes in C. elegans, B. malayiand Drosophila encode in the quality control of N-glycosylated proteins in homologous proteins that contain cysteine in place of ER mediated by the cysteine-rich domain of SELE- selenocysteine [2]. The NMR structure of fruit fly NOF. Thus, SELENOF could possibly be involved in glycoprotein folding control in ER [5, 6]. Tunicamycin (an N-linked glycosylation inhibitor and is commonly * Correspondence: [email protected] used to induce ER stress experimentally) upregulates 1College of Life Sciences and Oceanography, Shenzhen Key Laboratory of Marine Bioresources and Eco-environmental Science, Shenzhen University, the SELENOF protein level, whereas dithiothreitol Shenzhen 518060, China (DTT, an ER stressor which blocks formation of Full list of author information is available at the end of the article © The Author(s). 2018 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Tian et al. Nutrition & Metabolism (2018) 15:7 Page 2 of 9 disulfide bonds in the ER preventing xidative folding Methods of membrane and secretory proteins) promotes SELE- Amplification and mutagenesis of cDNA encoding NOF degradation through the proteasome pathway SELENO F and plasmid construction [7]. SELENOF knock out (KO) mice demonstrated no The cDNA encoding human SELENOF was cloned from obvious phenotypes, except the development of nu- a human fetal brain cDNA library (Clontech, U.S.A). The clear cataracts resulting from improper protein fold- primers, restriction enzymes and plasmids constructed for ing and posttranslational modifications that always this study are all presented in Table 1. Due to the absence induce ER stress [8]. The deficiency of SELENOF in of selenoprotein synthesis machinery in yeast cells, the Chang liver cells induces ER stress and inhibits cell Sec-coding TGA codons in SELENOF was changed into proliferation and invasion [9]. This outcome is con- Cys-coding TGC by using site-directed mutagenesis [20] sistent with the results of a knockdown of SELENOF to generate the SELENOF’ gene, which was then inserted mRNA in a colon cancer cell line, which resulted in into vector NpGBKT7 (Clontech, U.S.A), pEYFP-N1 the inhibition of tumor cell growth, and lung metas- (kindly provided by professor Deming Gou from Shen- tasis [10, 11]. Knockout of SELENOF in mice also zhen University) and PET28b (Novagen), to generate prevented the formation of chemically induced aber- NpGBKT7-Seleno F′, pECFP-C1-SELENOF,’ and SELE- rant crypts [12]. However, deletion of SELENOF in NOF’-PET28b for Yeast two-hybrid, FRET and pull down NIH3T3 cells led to cytoskeleton remodeling and experiments, respectively. The plasmids were confirmed membrane blebbing but not to apoptosis [13]. No ER to contain the target gene fragments by restriction enzyme stress was detected in the liver of SELENOF KO analysis and DNA sequencing. mice either [8]. The data imply that SELENOF plays multiple roles in different tissues or cells. SELENOF Library screening through yeast two-hybridization is discovered to distribute in brain with high level. It Plasmid NpGBKT7-SELENOF’ was used to screen the is explored from Atlas dataset that SELENOF gene human fetal brain cDNA library via the yeast two-hybrid expression is rich in neurons in olfactory bulb, system. Yeast transformation and library screening were hippocampus, cerebral cortex, and cerebellar cortex performed following the procedures described in the [14, 15]. It also has high transcript levels in mouse user manuals for the assays (YeastmakerTM Yeast neuronal cells of the hippocampus and Purkinje cells Transformation System 2 and Matchmaker™ Gold Yeast of the cerebellar cortex, with expression profiles that Two-hybrid System, Clontech, U.S.A). Positive yeast col- are similar to the ER chaperones calnexin (CNX) and onies from library screening were selected from the oxido-reductase ERp57, which are the major compo- quadruple dropout solid medium SD/−Ade/-His/−Leu/ nents of the quality control mechanism in ER [7], so −Trp containing X-α-Gal and Aba. Plasmids extracted SELENOF might play some roles in central neural from the yeast cells were transformed into E. coli Top10 system (CNS). competent cells. Prey plasmids were amplified in LB Human retinol dehydrogenase 11 RDH11 belongs to medium containing ampicillin and extracted from E. coli the short-chain dehydrogenases/ reductases (SDR) fam- cells. The obtained prey plasmids and the NpGBKT7- ily [16]. It is able to reduce both all-trans- and cis- SELENOF’ bait plasmid were co-transformed into retinaldehydes into all- trans- and cis- retinol (Vitamin A) [17, 18]. An RDH11 knockout in mice did not affect Table 1 Primers used for plasmids construction the retinoid profiles when the mice adapted to dark Plasmids Sequence conditions [17], but the deleterious mutations in the constructed RDH11 gene caused a new syndrome with retinitis pig- SEL F′-YFP-N1 F 5’-ATGGTAGCGATGGCGGCTGGGC-3′ mentosa [19], which is an inherited degenerative dis- R5’-TTATATGCGTTCCAACTTTTCAC-3′ ease in the retina that causes severe vision impairment. NpGBKT7-Sel F′ F5’-TTTGGGGCAGAGTTTTCATCGGAG-3’ Therefore, RDH11 may be related to not only the vita- R5’-TTATATGCGTTCCAACTTTTCAC-3’ min A cycle but also retina protection.To obtain insight RDH11-YFP-N1 F 5’-CGGAATTCATGGTTGAGCTCATGTTCCCGCTGT-3’ into the function of SELENOF in the brain, an inter- R5’-TAGGTACCCTGTCTATTGGGAGGCCCAGCAGGTC-3’ active protein of SELENOF was screened out to be RDH11 with a yeast two-hybrid system and validated RDH11-CFP-C1 F 5’-CGGAATTCCGTTGAGCTCATGTTCCCGCTGT-3’ with fluorescence resonance energy transfer (FRET), R5’-TAGGTACCTGGGTCCAACTGGCACTGCCTGTTA-3’ co-immunoprecipitation (co-IP) and pull-down assays SEPT3-CFP-C1 F 5’-ATAAGCTTATTCCAAAGGGCTCCCAGAGAC-3’ in this study. Overexpression of SELENOF in R5’-TAGGTACCTCATGGGTTACTGTCGTGGCTTT-3’ HEK293T cells inhibited enzyme activity of RDH11, TUBB4-CFP-C1 F 5’-TAGAATTCACGGGAGATCGTGCACCTGCAGG-3’ implying the function of SELENOF in retinol R5’-TAGGTACCCTAGGCCACCTCCTCCTCCGCCT-3’ metabolism. Tian et al. Nutrition & Metabolism (2018) 15:7 Page 3 of 9 Y2HGold yeast cells and grown on SD /−Leu /−Trp /x- N1, SELENOF-HA-SelExpress1, RDH11-pEYFP-N1 and α-gal /Aba solid medium to determine whether the col- HA-SelExpress1 were transfected into cells as the con- onies turned blue. The positive prey plasmids were then trol conditions. Forty eight hours after transfection, the sequenced and analyzed with the basic local alignment cells were lysed in RIPA solution