Horizontal Gene Transfer in the Evolution of Photosynthetic Eukaryotes

Journal of Systematics and Evolution 51 (1): 13–29 (2013) doi: 10.1111/j.1759-6831.2012.00237.x

Review Horizontal gene transfer in the evolution of photosynthetic eukaryotes 1,2Jinling HUANG* 1,2Jipei YUE 1(Department of Biology, East Carolina University, Greenville, North Carolina 27858, USA) 2(Key Laboratory of Plant Biodiversity and Biogeography, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, China)

Abstract Horizontal gene transfer (HGT) may not only create genome mosaicism, but also introduce evolutionary novelties to recipient organisms. HGT in plastid genomes, though relatively rare, still exists. HGT‐derived genes are particularly common in unicellular photosynthetic eukaryotes and they also occur in multicellular plants. In particular, ancient HGT events occurring during the early evolution of primary photosynthetic eukaryotes were probably frequent. There is clear evidence that anciently acquired genes played an important role in the establishment of primary plastids and in the transition of plants from aquatic to terrestrial environments. Although algal genes have often been used to infer historical plastids in plastid‐lacking eukaryotes, reliable approaches are needed to distinguish endosymbionts‐derived genes from those independently acquired from preferential feeding or other activities. Key words endosymbiosis, genome evolution, plants, plastids.

Gene transfer across the boundaries of species or In eukaryotes, gene transfer may occur between genomes has been widely recognized as an important genomes (mitochondrial, plastid, and nuclear) within force in the evolution of life on earth (Gogarten the cell or between distinct organisms. The former is et al., 2002; Andersson, 2005; Simonson et al., 2005; often referred to as intracellular gene transfer (IGT), Keeling & Palmer, 2008). According to Woese (2002), which is mostly unidirectional from mitochondria or the primary cellular evolution was communal and gene plastids to the nucleus and sometimes between plastids transfer was the driving force in the origin of modern and mitochondria. Because of the endosymbiotic origin cells. Thus far, an increasing amount of data shows that of mitochondria and plastids, this process is also called the impact of gene transfer extends far beyond the origin endosymbiotic gene transfer (EGT). The process of of cellular life. In prokaryotes, because mutations are genetic movement between species is termed horizontal mostly deleterious, acquisition of foreign genes is or lateral gene transfer (HGT or LGT). Although the considered the major means to gain novel genes. By role of HGT in eukaryotic evolution was poorly acquiring ready‐to‐use genes from other sources, gene appreciated historically, an increasing amount of data transfer may confer recipient organisms beneficial indicates that HGT not only is frequent in unicellular phenotypes and immediate abilities to explore new eukaryotes of various habitats and lifestyles (Andersson, resources and niches (Gogarten et al., 2002; Jain et al., 2005; Keeling & Palmer, 2008), but occurs in multi- 2003). Indeed, acquired genes are reportedly respon- cellular eukaryotes as well (Scholl et al., 2003; Pierce sible for the spread of photosynthesis, nitrogen fixation, et al., 2007; Moran & Jarvik, 2010; Ni et al., 2012; Yue bacterial pathogenesis, and antibiotic resistance. It has et al., 2012). There is also clear evidence that HGT been estimated that almost all prokaryotic genes have occurs between eukaryotic species (Rumpho et al., been affected by gene transfer in their evolution (Dagan 2008; Moran & Jarvik, 2010; Sun et al., 2010, 2011). In et al., 2008). As a result, gene transfer may not only many cases, acquisition of genes from other species has spread evolutionary success across distinct lineages, but significantly shaped the evolution of the biochemical also potentially transform the Darwinian tree‐like system of recipient organisms. phylogenesis pattern into a net over time (Doolittle, This article reviews the occurrence and impact of 2000; Bapteste et al., 2009; Martin, 2011). HGT in the evolution of photosynthetic eukaryotes. Given the availability of several reviews related to HGT in plant mitochondria (Richardson & Palmer, 2007; Keeling & Palmer, 2008), we will limit this review to Received: 15 January 2012 Accepted: 4 November 2012 * Author for correspondence. E‐mail: [email protected]. Tel.: 252‐328‐ HGT in plastid and nuclear genomes of photosynthetic 5623. Fax: 252‐328‐4178. eukaryotes.

1 Detecting horizontally acquired genes in related taxa will not be known. This in turn can eukaryotes translate into difficulties distinguishing recent HGT versus ancient HGT. A good and practical Because gene transfer involves the origin of a gene approach should be a broad and balanced from another genome or species, any evidence taxonomic sample from representative groups indicative of evolutionary relationships can be used of all three domains of life (bacteria, archaea, to identify transferred genes. Such evidence, for and eukaryotes). In many cases, identification of example, may include GC content or codon usage the exact donor involves re‐sampling once the patterns that often differ between donor and recipient donor has been narrowed down. organisms. Since phylogenetic analyses investigate the (2) Organismal phylogeny. A good knowledge of relationships among sampled sequences, they are often organismal phylogeny is not only essential for considered to be the most reliable method to identify the identification of transferred genes, but also transferred genes (Andersson, 2005; Keeling & particularly important for explaining some Palmer, 2008). Theoretically, if a gene in a eukaryotic puzzling gene phylogenies resulting from differ- genome was acquired from a bacterium, it is essentially ential displacement (Huang & Gogarten, 2009). a bacterial gene in origin and should be more closely The scenario arising from differential displace- related to bacterial homologs than to other eukaryotic ment of preexisting homologs by acquired genes sequences. Therefore, evidence for gene transfer can be is potentially frequent, but largely ignored in inferred by comparing the gene phylogeny and the current studies and rarely discussed in literatures. known organismal phylogeny for incongruence. Such a scenario may have been frequently Though straightforward as it sounds, the phylo- mistaken as resulting from independent acquis- genetic approach can be subject to many potential itions, or gene duplication followed by differ- pitfalls due to data quality, methodology, differential ential losses. gene gain/loss, or other issues (Huang & Gogarten, (3) Gene distribution. Accurate information of gene 2006). In particular, long‐branch attraction due to distribution can eliminate some spurious cases unequal sequence changing rates can lead to erroneous of gene transfer, but also is extremely useful for gene tree topologies. Biased or insufficient taxonomic distinguishing horizontally acquired genes from sampling may also result in grouping unrelated those arising from other scenarios, including sequences in a clade, which is superficially similar to differential losses and IGT events. For example, cases of gene transfer. Even if the gene tree is free of a eukaryotic gene that is absent from cyanobac- artifacts, other explanations always exist. In particular, teria and proteobacteria may not have a plastid differential gene loss, sometimes associated with or mitochondrial origin. The determination of duplication and paralogy, can always be invoked as gene distribution entails extensive searches of an explanation alternative to HGT (Gogarten & many additional databases other than GenBank. Townsend, 2005; Huang & Gogarten, 2006). This (4) Gene neighborhood. Some acquired genes are alternative scenario, however, becomes increasingly functionally related, but their exact donors are less likely if more and more loss events need to be often uncertain. However, if these genes are invoked. In some other cases, independent or recurrent physically adjacent or form an operon on a gene transfer events may also lead to gene tree chromosome of the potential donor, a likely topologies that are difficult to interpret. scenario would be that these genes were To effectively reduce the number of false positives acquired together from the same donor (Ricard in HGT detection, a combined approach including et al., 2006; Sun et al., 2010). multiple lines of independent evidence is preferred. The (5) Pairwise sequence comparison. Although se- following summarizes some practical considerations in quence similarity comparison is not an accurate the identification of acquired genes in eukaryotes. approach to detect transferred genes, its sensi- tivity is high (Frickey & Lupas, 2004). For (1) Broad and balanced taxonomic sampling.A example, the vast majority of plastid genes broad and balanced sampling is not only either are highly similar to cyanobacterial important for narrowing down the donor, but homologs in pairwise sequence comparison the recipient as well. If the donor taxon is not or group with cyanobacterial homologs in sampled, it cannot be identified. On the other phylogenetic analyses (Huang J, unpublished hand, if the recipient group is not properly data). Therefore, if a eukaryotic sequence sampled, the distribution of an acquired gene in is most similar to their cyanobacterial or a‐

proteobacterial homologs, an organellar origin primary plastids evolved from a cyanobacterial cell of this gene cannot be excluded. engulfed by an early eukaryote. The exact systematic (6) Timing of emergence for HGT partners. This is position of the progenitor of primary plastids within important for pinpointing the donor and the extant cyanobacteria remains less certain (Deusch recipient for transfer partners. The donor must et al., 2008; Criscuolo & Gribaldo, 2011), but there is emerge at least as early as the recipient (Huang evidence that this progenitor, like other free‐living & Gogarten, 2006). cyanobacteria, was subject to frequent HGT (Zhax- (7) Rare genomic or molecular characters.Exam- ybayeva et al., 2006; Gross et al., 2008). Such a ples include gene fusion, domain structures, cyanobacterial cell was enslaved and eventually indels, and other rare genomic characters. domesticated into primary plastids that set the stage Sharing these rare characters often is an for eukaryotic photosynthesis. The primary plastids are indication of potential gene transfer (Sun still found in red algae, green plants and glaucophytes, et al., 2010; Yue et al., 2012). which are collectively called Plantae or primary photo- (8) Genomic scaffold information.Manygenome synthetic eukaryotes. These primary plastids were sequencing projects are contaminated by bacte- further spread via secondary or higher‐level endo- rial sequences (e.g., by associated bacterial symbioses into multiple other major eukaryotic lineages endosymbionts). These bacterial sequences (or such as alveolates, diatoms, haptophytes, cryptophytes, genes) sometimes are included in genome and euglenids (Delwiche, 1999; Archibald, 2009; annotation and may be mistaken as HGT‐ Keeling, 2010; Katz, 2012; Fig. 1). Additionally, derived. However, because of their foreign independently evolved cyanobacterial endosymbionts nature, these sequences are often located on in various stages of integration have also been found in small contigs or scaffolds consisting of a few other eukaryotes (Prechtl et al., 2004; Marin et al., bacterial genes. Therefore, genomic scaffold 2005; Bodyl et al., 2007). In particular, the amoeboid information is often needed to eliminate the Paulinella chromatophora contains cyanobacterial possibility of sequence contamination (Sun & endosymbionts (chromatophores) that are related to Huang, 2011; Yue et al., 2012). Prochlorococcus and Synechococcus. Such independ- (9) Artifact control. Some genomic or EST data ently evolved chromatophores can be stably inherited contain many artifacts due to contamination. and their genomes are reduced, with many of their genes Identification of the same gene in multiple transferred to the host nucleus (Nowack et al., 2008, sources (ESTs, genomic sequences, wet‐lab 2011; Reyes‐Prieto et al., 2010). experiments etc.) is a useful way to ascertain its Genome reduction as a result of IGT and gene loss existence in the genome (Huang et al., 2004a). epitomizes the integration and transformation of an (10) Phylogenetic resolution. Because of the required endosymbiont into an organelle (Martin & Herrmann, phylogenetic resolution varies among taxonomic 1998; Selosse et al., 2001). During the process of ranks, lack of sufficient resolution or presence of organellogenesis, dispensable genes were lost, whereas phylogenetic artifacts at lower taxonomic ranks many other genes were transferred to the nucleus of the may not affect the identification of gene transfer host cell (Doolittle, 1998; Martin & Herrmann, 1998). events between groups of higher ranks (Woese, Gene transfer from organelles to the nucleus was also 2000; Huang & Gogarten, 2009). A gene accompanied by the evolution of an organelle‐specific phylogeny may rarely reflect “perfect” organ- protein translocation machinery (Jarvis, 2008), as ismal relationships within each domain of life recently demonstrated in P. chromatophora (Bodyl (bacteria, archaea, or eukaryotes), but this should et al., 2010b; Mackiewicz et al., 2012; Nowack & not significantly affect the identification of inter‐ Grossman, 2012). Eventually, the protein products for domain transfer events. many of the IGT‐derived genes are directed by an N‐terminal targeting signal and imported back to the original organelles, though the proteins for some others 2 Endosymbioses and eukaryotic may function in the cytosol (Martin & Herrmann, 1998; photosynthesis Martin, 2010). IGT appears to be a dynamic process and sometimes occurs in large fragments. The proportion of The origin of plastids represents one of the most nuclear genes that are derived from plastids has been significant events in eukaryotic evolution and high- estimated from 10% to 25% in some photosynthetic lights the importance of organismal interaction and eukaryotes (Reyes‐Prieto et al., 2006; Deusch et al., integration in macroevolution. It is now clear that 2008). Such frequent IGT from plastids also prompted a

Serial secondary endosymbiosis Tertiary endosymbiosis

m m N N N m

Lepidodinium Kryptoperidinium karlodinium

Secondary endosymbiosis

m N m N N m N m N m N m N m

Euglenids Chlorarachniophytes Dinoflagellates Apicomplexans Heterokonts Haptophytes Cryptophytes

m N m C N P Land plants Green algae Red algae

m m Primary endosymbiosis N N C C Glaucophytes

m N

+ N m

Cyanobacteria Eukaryotes

Fig. 1. The origin and spread of plastids based on current knowledge of eukaryotic evolution. It is generally agreed that plastids evolved from a single primary endosymbiotic event in which an early heterotrophic eukaryote engulfed a cyanobacterium. This primary endosymbiosis gave rise to three extant photosynthetic lineages (glaucophytes, red algae, and green plants). The red algal plastid was further spread to cryptophytes, haptophytes, heterokonts, apicomplexans, dinoflagellates, and others through either secondary or tertiary endosymbioses. Euglenids and chlorarachniophytes are thought to have acquired their plastids from green algae in two separate secondary endosymbiotic events. Within the dinoflagellates Kryptoperidinium, Karlodinium, and Lepidodinium, the ancestral plastid has been replaced by primary or secondary plastids through tertiary endosymbioses or serial secondary endosymbioses. Dashed lines show uncertain evolutionary events. Please note that this scheme is only for demonstrating series of endosymbioses among extant eukaryotic lineages. Scheme was drawn based on Bodyl et al. (2009), Archibald (2009), and Keeling (2010). hypothesis that primary photosynthetic eukaryotes may IGT is limited: the source of transferred genes should be not be closely related and their relationships may have no greater than the genome of the a‐proteobacterial resulted from convergent evolution due to IGT from progenitor of mitochondria and/or the cyanobacterial plastids (Stiller, 2007). progenitor of plastids. Because of the gradual genome The frequent IGT also leads to difficulties in reduction over time for both mitochondria and plastids, identifying HGT‐derived genes. To identify any gene IGT will gradually decrease. This process may even- independently acquired from bacteria, it is necessary tually stop if organellar genomes are reduced to the to exclude the possibility that the gene is actually of extreme or become entirely lost (e.g., in the apicom- mitochondrial or plastid origin. Although genes acquired plexan Cryptosporidium). On the other hand, if these through both IGT and HGT are ultimately of foreign organelles are not intrinsically resistant to transfer origin, the former are derived from a few endosymbionts, events, genes from extra‐genomic sources may be whereas the latter were acquired from diverse organisms potentially accumulated in them. This is particularly by independent events. This distinction allows us to evident for plant mitochondria, which have acquired understand the contributions of different genetic sources genes from miscellaneous sources (Richardson & and mechanisms to eukaryotic evolution. Palmer, 2007).

3.1 Gene transfer to plastid genomes 3 HGT in plastids Although gene transfer from plastids to the nucleus and mitochondria are frequently reported in plastid‐ Physical contact or adjacency often facilitates gene containing eukaryotes, the reverse is rarely known. In transfer (Jain et al., 2003). Despite the favorable particular, no transfer event has been reported from the intracellular environment for transfer between organ- nucleus to plastids to our knowledge, and only a few elles or from organelles to the nucleus, the gene pool for cases are known from mitochondria to plastids. It is

© 2012 Institute of Botany, Chinese Academy of Sciences HUANG & YUE: HGT in plants 17 important to note that, although several mitochondria‐ 3.2 Gene replacement versus acquisition of novel derived genes are discussed here, it is not entirely clear genes whether they were transferred from within the cell or Acquired genes can be functionally classified into other organisms. The plastid genome of Daucus carota homologous replacement and introduction of novel contains a DNA fragment of 1,439 bp that is not found functions. A strict homologous replacement may in any other plastid genomes, but homologous to contribute to genome mosaicism, but not phenotypic mitochondrial chromosomal regions of flowering plants changes. Homologous replacement often is involved in (Goremykin et al., 2009). Similar scenarios were found functions that are essential for all organisms or that for genes encoding a putative DNA polymerase type B define a major lineage. In plastid‐containing eukar- (dpoB) and a tyrosine recombinase family protein (int) yotes, these gene replacement events involve ribosomal in plastid genomes of the green alga Oedogonium proteins in apicomplexans, cryptophytes and hapto- cardiacum and the heterokont Heterosigma akashiwo, phytes, as well as the rbcL operon in red algae. The respectively (Brouard et al., 2008; Cattolico et al., dnaX gene in cryptophytes, which presumably encodes 2008). the gamma/tau component of DNA polymerase, was Another case of transfer from mitochondria to likely acquired from a firmicute bacterium (Khan plastids involves the protozoan group apicomplexans, et al., 2007). Because homologous sequences of dnaX which are mostly known as parasitic pathogens (e.g., are found in cyanobacteria, the acquisition of a malaria parasite Plasmodium). Apicomplexans contain firmicute‐like dnaX may represent either replacement some of the smallest mitochondrial genomes known of preexisting plastid homologs, or gene acquisition to date, but also often a relict and non‐photosynthetic following the loss of the plastid copy. Interestingly, the plastid (apicoplast) that is derived from an algal dnaX homologs in land plants are about 800 amino endosymbiont. The mitochondrial genomes of acids (aa) longer than bacterial copies and involved in Plasmodium, Theileria,andBabesia are about 6 kbp regulating trichome development (Ilgenfritz et al., containing only three protein‐coding genes, whereas 2003). If these land plant dnaX homologs are of plastid Cryptosporidium appears to have lost its entire origin (they form a monophyletic group with cyano- mitochondrial genome. Despite the ultimately cyanobacterial sequences with modest support), it will point bacterial origin of apicoplasts, earlier studies suggest that to the contribution of plastid gene transfer to the three apicoplast ribosomal protein genes are likely evolution of new genes. derived from mitochondria (Obornik et al., 2002). At least two acquired genes (dpoB and int) are In fact, molecular phylogeny consistent with this earlier novel to their recipient plastid genomes, both of which study is provided from analyses of each of the apico- are likely of mitochondrial ancestry (Brouard et al., plast ribosomal proteins (Huang J, unpublished data). 2008). Whether these genes are functional is not Because ribosomal protein genes often form an operon in entirely clear, but both genes share conserved regions bacteria, the replacement of plastid copies by mitochon- characteristic of their respective gene families. It has drial homologs likely occurred in a large DNA fragment. been suggested that, even if dpoB is functional, its role Several HGT cases from other bacteria to plastids may be marginal due to the lack of a dedicated DNA also are known (Delwiche & Palmer, 1996; Rice & replication machinery in plastids (Brouard et al., 2008). Palmer, 2006; Khan et al., 2007; Janouskovec Because int is often found in bacteriophages and et al., 2010; Moszczynski et al., 2012). The Rubisco plasmids and it is responsible for excision and inte- operon of red algae was likely acquired from gration of DNA fragments, its transfer among genomes proteobacteria (the two component genes, rbcL and is not entirely unexpected. rbcS, are separated by only a few base pairs on chromosomes of some proteobacteria; Delwiche & 3.3 Why are transferred genes relatively rare in Palmer, 1996). The ribosomal rpl36 gene was found to plastids? have been transferred from bacteria to haptophytes and It is often believed that genome complexity cryptophytes and interpreted as evidence for their constitutes a major barrier to gene transfer and that monophyly (Rice & Palmer, 2006), though more recent genes whose products are involved in complex cellular analyses suggest that these two groups may not be interactions, such as transcriptional and translational closely related (Burki et al., 2012). Evidence for HGT machineries, are often resistant to gene transfer (Jain from bacteroidetes to plastid minicircles in dinoflagel- et al., 2002; Ciccarelli et al., 2006). This notion not only lates has also been recently reported (Moszczynski explains the observation that the vast majority of et al., 2012). All these reported HGT events occurred in transferred genes are functionally related to metabo- algae. lisms, but also underpins the long‐standing practice of

© 2012 Institute of Botany, Chinese Academy of Sciences 18 Journal of Systematics and Evolution Vol. 51 No. 1 2013 using 16S RNAs and, occasionally, ribosomal proteins, 2012), the vast majority of acquired genes may for reconstructing organismal relationships. Never- represent homologous replacement of these genes. A theless, replacement of several genes encoding riboso- similar scenario has also been observed in plant mal proteins has been found in plastids. In addition, mitochondria, where most of the acquired genes numerous inteins, endonucleases, and other selfish encode ribosomal proteins or are related to respiration elements associated with Group I and Group II introns (Richardson & Palmer, 2007). This may partially exist in plastid genomes (Turmel et al., 2002; Pombert explain the observation that homologous replacement et al., 2005; Khan & Archibald, 2008; Smith & of ribosomal proteins occurred multiple times in plastid Lee, 2009). This suggests that, even though the total genomes (Obornik et al., 2002; Rice & Palmer, 2006). number of acquired genes in plastid genomes is lower, While the transcriptional and translational machineries plastids are not intrinsically resistant to gene transfer. are universal to all organisms, the capability of photo- Then, what leads to the seemingly lower number of synthesis, on the other hand, exists only in a restricted acquired genes in plastids? number of prokaryotic groups other than cyanobacteria It is possible that the lower number of acquired (Xiong, 2006). Such restricted taxonomic distribution genes is related to the endosymbiotic nature of plastids of photosynthesis may limit the potential of homolo- and their reduced genomes. Such genome reduction has gous replacement for photosynthesis‐related genes. often been attributed to multiple, sometimes related, This does not diminish the possibility of homologous causes. The active electron transport in bioenergetic replacement from other cyanobacterial taxa, but cases organelles (plastids and mitochondria) can lead to a high of gene replacement between closely related taxa, concentration of oxygen free radicals, which are highly though most frequent (Raymond et al., 2002), are often toxic to the cell and mutagenic to DNA molecules. This difficult to distinguish from phylogenetic artifacts. increased mutagenic load may have driven the trans- Compared to plastids, gene transfer is relatively location of organellar genes to the nucleus (Allen & frequent in mitochondria, in particular for seed plants Raven, 1996; Allen et al., 2005). Another possible (Richardson & Palmer, 2007; Goremykin et al., 2009). cause is the effect of Muller’s ratchet (Blanchard & Not only do plant mitochondrial genomes contain genes Lynch, 2000). Plastids are haploids and uniparentally acquired from plastids and viruses, there are also inherited, thus having smaller effective population sizes. multiple reports of HGT between mitochondria of They also exist in an isolated intracellular environment distantly related species (Richardson & Palmer, 2007; and often lack recombination. Therefore, deleterious Mower et al., 2010; Sanchez‐Puerta et al., 2011). In mutations tend to accumulate in plastid genes because of particular, the mitochondria of Amborella, a basal the effect of genetic drift (Martin & Herrmann, 1998), angiosperm, appear to contain a considerable number of which may further lead to gene transfer to the nucleus genes that were acquired from mitochondria of other and plastid genome reduction. However, because plants (Bergthorsson et al., 2004; but also see genome reduction also occurs in algal endosymbionts Goremykin et al., 2009). The frequent gene transfer (nucleomorphs) that do have active redox activities and between seed plant mitochondria is often linked to because recombination does occur in organelles of some mitochondrial fusion and might have been facilitated by organisms, selection for efficient replication and physical contact through epiphytes and parasitic plants metabolism may also account for the reduction of (Bergthorsson et al., 2004; Rice & Palmer, 2006). organellar genomes (Selosse et al., 2001). Although fusion events also have been reported on Genome reduction may at least partially explain plastids and mitochondria of other eukaryotic groups the rarity of novel gene acquisition in plastids. For (Cavalier‐Smith, 1970; Kavanagh et al., 1999), the functional plastids, only genes responsible for redox mitochondria of land plants are larger and dynamic due regulation may ultimately be retained (Allen et al., to frequent duplication and recombination (Palmer 2005). Such reduced plastid genomes with a dedicated et al., 2000), which may potentially alleviate the effects gene set may decrease their potential to accommodate of Muller’s ratchet and allow accommodating foreign other genes with nonessential functions. Unless other genes (Selosse et al., 2001). acquired DNA molecules can mobilize to other chromosomal locations (e.g., inteins or intron‐associated endonucleases) or to the nucleus, they will likely 4 Frequent HGT to nuclear genomes decay into pseudogenes over time and be eliminated from plastids. Additionally, because most genes 4.1 Primary photosynthetic eukaryotes encoded in plastids are essential for translation and Hybridization among plant species is frequent in photosynthesis (Race et al., 1999; Dorrell & Howe, nature and they are routinely used in horticultural and

© 2012 Institute of Botany, Chinese Academy of Sciences HUANG & YUE: HGT in plants 19 crop breeding to achieve desirable traits (Rieseberg & Most HGT‐derived genes identified in primary Carney, 1998; Soltis & Soltis, 2009). Introgressive photosynthetic eukaryotes were more anciently ac- hybridization, which occurs through repeated back- quired by the ancestral lineages (Copley & Dhillon, crossing and causes gene movement from one parental 2002; Huang & Gogarten, 2006, 2007, 2008; Emiliani species to another, essentially represents HGT between et al., 2009; Richards et al., 2009; Price et al., 2012). In closely related species. Because plant genomes are particular, at least 38 gene families were transferred relatively stable in gene content, introgressive hybrid- from other organisms to the ancestors of green or land ization in plants rarely involve novel genes. Such plants (Yue et al., 2012) and some others were to the interspecific HGT appears to have a limited impact on ancestor of primary photosynthetic eukaryotes (Huang the origin of major plant lineages. & Gogarten, 2008; Price et al., 2012). These data are Like in other eukaryotes, a perceivable source of consistent with the suggestion that foreign genes foreign genes for primary photosynthetic eukaryotes acquired by unicellular ancestors of multicellular includes distantly related taxa, in particular bacteria eukaryotes may be accumulated in descendant lineages (Andersson et al., 2003; Loftus et al., 2005; Ricard over time (Huang & Gogarten, 2006). Although most of et al., 2006). Such distant donors can be identified more these anciently acquired genes were from prokaryotes, confidently using the phylogenetic approach and they at least 11 genes were acquired from fungi (Richards are also more likely to introduce novel genes and et al., 2009; Yue et al., 2012), presumably during the phenotypes. Because HGT is a dynamic process conquest of land by early green plants. Because these occurring over time, foreign genes might have been genes have been retained in plants over a long period of introduced anciently to the ancestors of major lineages time, many of them are important for plant develop- or more recently to individual organisms. Thus far, ment, regulation, and adaptation. complete genomes of major green plant lineages are An interesting example of HGT in primary available, and those for the red alga Cyanidioschyzon photosynthetic eukaryotes involves the bacterial group merolae and the glaucophyte Cyanophora paradoxa chlamydiae (Horn et al., 2004; Huang & Gogarten, have also been published (Matsuzaki et al., 2004; Price 2007). Members of chlamydiae are commonly found as et al., 2012). These sequence data offer an opportunity obligate endosymbionts in animals and environmental to assess the dynamics and importance of HGT in the amoebae, but no chlamydial species has been identified evolution of primary photosynthetic eukaryotes. in plants and other plastid‐containing eukaryotes. Several cases of recent HGT have been reported in Several earlier genome analyses identified an unex- primary photosynthetic eukaryotes. One of these pected number of chlamydial genes that are most reported cases involves a gene of unknown function similar to plant sequences, most of which are also that was transferred from a Sorghum‐related species to plastid‐targeted. This observation led to several hypo- the parasitic plant Striga hermonthica (Yoshida theses involving various evolutionary scenarios, in- et al., 2010). It has been speculated that such a transfer cluding shared ancestry between chlamydiae and was in the form of mRNA and mediated via haustoria of cyanobacteria (plastids), HGT from plant‐like organ- S. hermonthica that connect to the host plant. Another isms to chlamydiae and vice versa (Stephens et al., case involves the grass Alloteropsis, which was 1998; Brinkman et al., 2002; Ortutay et al., 2003). suggested to have acquired C4 photosynthesis at least Further phylogenomic analyses of photosynthetic four times in the past 10 million years (Christin eukaryotes clearly indicated that these genes, including et al., 2012). Recently acquired genes have also been multiple transporter genes, were acquired from chla- identified in nonvascular plants such as the moss mydiae (Huang & Gogarten, 2007; Tyra et al., 2007; Physcomitrella patens (Yue et al., 2012) and the Becker et al., 2008; Moustafa et al., 2008; Collingro lycophyte Selaginella moellendorffii (Huang J & Yue J, et al., 2011; Price et al., 2012). Based on the distribution unpublished data). Other than the above examples, of chlamydiae‐related genes in primary photosynthetic cases of recent HGT reported in primary photosynthetic eukaryotes and the fact that all extant chlamydial eukaryotes are relatively rare in general. The sparse species are obligate endosymbionts, chlamydial genes report of recently acquired genes, however, by no in plants most likely resulted from an ancient chlamy- means should be interpreted as evidence of lacking dial endosymbiont in the ancestor of primary photo- recent HGT in primary photosynthetic eukaryotes. In synthetic eukaryotes. These acquired chlamydial genes fact, some casual sequence similarity search will might have facilitated the transition of cyanobacterial identify genes specific to these organisms and endosymbionts into primary plastids (Huang & prokaryotes, which usually suggest gene transfer events Gogarten, 2007; Moustafa et al., 2008; Price et al., (Figs. 2 and 3). 2012).

Cyanidioschyzon L CV HGI DA I L QY I K K CY GS L F S P RA I A Y RE E K GY P HL DMRL S V GV MQMV QS R- - V S GV CF T L DP V T GK RDL I V I E CCWGL 224 Ammonifex L NV RGT A E V V E A V RK CWA S L WT A RA T A Y RQA QGF DHF A V Y L S A V V QK MI QS E - - RS GV A F T V NP V T GV RDE L MI NA S WGL 213 Aminobacterium L HI RGA E E V L K HV QK CWA S L WT A RA T Y Y RE K QDF NHF E V A L S A V V QK MV RS E - - K S GV L F T A NP V S NDL NQI MI NA S WGL 216 Sulfolobus L NV S - K GE L L DA V K K V WA S L Y T A RA I S Y RRF K GI DQI T V E MA V V V QK MV NS R- - S A GV MF T L HP V T GDRNY I MI E S S WGL 203 Metallosphaera L NV N- RS NI I E A V K L V WA S L Y NE RA I E Y RK S K GI DS S K V E MA V V V QK MV NS K - - S S GV MF T L NP S NGDRNL I V I E S S WGL 211 Desulfurococcus L NV Y GA DS V V Y HV K RCWA S L F T A RA T F Y RV A QGI P HE K T F MS V T V QK MV NS R- - S A GV MF T L HP V T GDE NV V V I E GS WGL 219 Pyrolobus L NV RGE DE V V K K V K A CWA S L F NA RA I F Y RE QMK I P HE K T Y MA V V V QK MV MS R- - S A GV MF T V HP V T GDDNV V V I E S A WGL 217 Lactobcillus L NV HGRDNV I QK V K E CY A S T F T DRA T Y Y RV K QGF P HE K V A L S A A V QMMA F S K - - A S GV MF T V NV A T GDDS K V MI E GS WGL 220 Megasphaera L NV QGA DT V I QK V K E CY A S T F T DRA T Y Y RE K K GF DHMDV A L S A A I QMMV F S K - - A A GV MF T V DL V T GDDS T I T I E GA WGL 219 Nostoc L NV HGL QA V L E S CHK CF A S I F T DRA I S Y RQI K GF DHF NI A L S V GV QK MV RS DL A T S GV MF S I DT E T GF K DA V L I T A A Y GL 237 Microcoleus L NV QA CA GV L E CCHK CF A S I F T DRA I S Y RQQRGF DHF E V A L S V GV QK MV RS DL A S S GV MF S I DT E T GF K NA V L V T A A Y GL 237 Arthrospira L NI HGI K GI L E A T RK CF A S L F T DRA I S Y RS L QGF DHF DV A L S I GV QK MV RS DL A T S GV MF S I DT E T GF K NA V I I T A A Y GL 235 Planctomyces L NV RGP A T L L E T CRRCF A S L F T DRA I S Y RT E K GF DHF DI A L S I GI QRMV RS DL A A S GV MF S I DT E T GF K DA V L I NA A Y GL 250 Caminibacter L NI K GE E NL L L A Y K MCL A S NF T DRS I S Y K Y T HNF DP L K V Y L S V V I MK MV RS DL A S S GV MF S I DT E S GF RDV V F I NA A Y GL 220 Methylacidiphilum L NI RGRDNL L S S I K K CF A S L F T DRA I S Y RT DMGF E HT K V A L S V GV QRMV RS DL A CS GV MF S I DT E S GF NNV V L I NS S Y GL 232 Proteus L NV QGI DA I L V A I K HV F A S L F NDRA I S Y RV HQGY DHRGV A L S A GV QRMV RS DL A S S GV MF T I DT E S GF DQV V F I T S A Y GL 226 Thalassiosira L NV V GK HS V CS A V L E CL A S V F T DRA I A Y RV HNGF DHMQV K GA V S V QL MV RS DL A S S GV A F T L DP DT GF RDV V V I T GS Y GL 276 Methanohalophilus L NI HGY HA L NDA CS RCF A S L F T DRA I S Y RI NNRF DHF DV GL S I GI MK MV RS DL A S S GV MF T I DT E T GF E DV V F I T GS Y GL 223 Xenopus L NV A GI A DV L HK MK E V F A S L Y NDRA I S Y RV HK GF A HDV V A L S A GV QRMV RS DL GA A GV MF T I DT E S GF E DV V F I T S S Y GL 225 Thermovibrio L NV RGI E NL L NS V K K CF A S L F T DRA I S Y RE T F GF DHF K V A L S V GV QK MV RS DL A S S GV GF S L DT E S GF K DV V L V T GS Y GL 224 Pseudomonas L NI RGV DNV I RA A K E V F A S L F NDRA I A Y RV HQGF DHK L V A L S A GV QRMV RS E T GT A GV MF T L DT E S GF RDV V F I T GA Y GL 220 Rhodanobacter L NV V GI DDV L HK V K E V F A S L Y NDRA I A Y RV HQGF K HE DV F L S A GV QL MV RS DV GA S GV L F T L DT E S GF RDV V F V T GS Y GL 222

B 80/62 Pyrolobus 100/100 Metallosphaera 98/98 Crenarchaeotes Sulfolobus 74/63 Desulfurococcus

73/92 100/100 Aminobacterium Synergistia Ammonifex Firmicutes 100/100 Cyanidioschyzon Red algae 100/100 Megasphaera Firmicutes Lactobacillus

59/56 Methanohalophilus Euryarchaeotes 100/100 Caminibacter Epsilon-proteobacteria Mariprofundus Zeta-proteobacteria Methylacidiphilum Verrucomicrobia Planctomyces Planctomycetes Campylobacterales Epsilon-proteobacteria 83/68 Microcoleus 99/100 Arthrospira Cyanobacteria Nostoc 77/93 Rhodanobacter */57 Pseudomonas Gamma-proteobacteria 99/100 Proteus 97/96 Lautropia Beta-proteobacteria 1 Xenopus Metazoa Thermovibrio Aquificales Novosphingobium Alpha-proteobacteria Simkania Chlamydias Aspergillus Ascomycetes Pelobacter Delta-proteobacteria Thalassiosira Diatoms Rubrobacter Actinobacteria

0.3

Fig. 2. Multiple sequence alignment (A) and molecular phylogeny of phosphoenolpyruvate synthase (B). Numbers above branches show bootstrap support values from maximum likelihood and distance analyses, respectively. The asterisk indicates value lower than 50%. The phosphoenolpyruvate synthase of the red algal Cyanidioschyzon merolae is most closely related to homologs from ﬁrmicutes and crenarchaeotes, and their relationship is also supported by several conserved amino acid residues and shared indels.

4.2 HGT in secondary and tertiary photosynthetic organisms (e.g., apicomplexans) also are important eukaryotes parasitic pathogens and their genomes have been Most secondary and tertiary photosynthetic or heavily sequenced. Because of their absence from plastid‐containing eukaryotes are unicellular and humans or distant relatedness to human homologs, presumably subject to frequent HGT. Some of these genes transferred from bacteria or plants to these

100/100 Bacillus Firmicutes Genome analyses for genes acquired from other Paenibacillus sources have also been reported on diatoms, brown 91/72 99/93 Pseudoxanthomonas Gamma-proteobacteria 69/73 Xanthomonas algae, and their close relatives oomycetes (Tyler 74/72 94/61 Asticcacaulis et al., 2006; Bowler et al., 2008; Cock et al., 2010). 100/100 Sphingomonas Alpha-proteobacteria In particular, 587 of the 10 402 (5.64%) genes in the 100/98 Novosphingobium Acidocella Alpha-proteobacteria pennate diatom Phaeodactylum tricornutum were */53 100/100 Volvox found to be of bacterial origin, over 300 of which are Green algae Chlamydomonas shared with the marine centric diatom Thalassiosira 72/87 Stigmatella Delta-proteobacteria pseudonana (Bowler et al., 2008). Many of the bacterial Parachlamydia Chlamydias Pseudomonas Gamma-proteobacteria genes in diatoms are related to nitrogen and carbon alpha proteobacterium BAL199 fixation, polyamine metabolism, and other important 100/100 Rhizobium Alpha-proteobacteria biochemical activities. In the oomycete plant pathogen Agrobacterium Phytophthora, which presumably is of photosynthetic 66/69 Mycobacterium Actinobacteria ancestry like the apicomplexan Cryptosporidium, many */52 Microbacterium Ketogulonicigenium Alpha-proteobacteria genes related to cyanobacterial and plant sequences Synechococcus Cyanobacteria were also identified (Tyler et al., 2006). In addition, Roseomonas Alpha-proteobacteria large viral DNA fragments or even complete virus‐like 91/91 Anabaena Cyanobacteria Agrobacterium Alpha-proteobacteria metabolic pathways were also found in brown algae, Dickeya haptophytes, and other phytoplanktons (Monier et al., Gamma-proteobacteria Pantoea 2009, 2011; Cock et al., 2010), though the transfer Roseomonas direction remains uncertain in some cases (Monier Agrobacterium Alpha-proteobacteria Ketogulonicigenium et al., 2009). Rhizobium Alpha-proteobacteria */54 Many other studies related to HGT in secondary 98/98 Acidovorax and tertiary photosynthetic eukaryotes were based on 59/82 Variovorax Beta-proteobacteria 100/99 Ralstonia EST data or analyses of individual metabolic pathways Burkholderia (Archibald et al., 2003; Obornik & Green, 2005; */63 Octadecabacter Alpha-proteobacteria Nosenko et al., 2006; Frommolt et al., 2008; Maruyama alpha proteobacterium BAL199 et al., 2011). These analyses indicate that both plastid 0.3 and nuclear proteomes in these organisms might be mosaic. Intriguingly, although it is often suggested that Fig. 3. Molecular phylogeny of oxidoreductase. Numbers above branches show bootstrap support values from maximum likelihood plastids of diatoms, haptophytes, cryptophytes, and and distance analyses, respectively. The asterisks indicate value lower other photosynthetic members of the controversial than 50%. Please note that the identifiable homologs of green algal group “chromalveolates” are derived from a red algal oxidoreductase (Volvox carteri gi|302829777; Chlamydomonas rein- hardtii gi|159474786) are only distributed in bacteria. endosymbiont (Keeling, 2009), a considerable number of sequences from these organisms showed strong pathogens may include some potential chemotherapeu- green algal affinities. Such phylogenetic signal from tic targets. This spurred some earlier phylogenomic green algae led to a speculation that these organisms analyses to search for acquired genes in apicomplexan might contain both red and green algal endosymbionts pathogens, which identified genes derived from both during their earlier evolution (Frommolt et al., 2008; the apicoplast organelle and prokaryotes (Abrahamsen Moustafa et al., 2009). et al., 2004; Huang et al., 2004a, 2004b). In the plastid‐ Some other eukaryotes do not have permanent lacking Cryptosporidium, genes that are related to plastids, but may harvest them through preying on algae cyanobacterial sequences and apicoplast/plastid pre- (kleptoplasty). Notably many sacoglossan sea slugs cursors were also identified (Huang et al., 2004a). These acquire transient plastids from their food algae and data support the hypothesis of plastid loss in retain them in their digestive gland cells. These Cryptosporidium (Lang‐Unnasch et al., 1998) and are harvested plastids (kleptoplasts) can perform photo- consistent with the later discovery of Chromera velia,a synthesis for various lengths of time, sometimes up photosynthetic alveolate closely related to apicomplex- to 14 months (Rumpho et al., 2000; Handeler et al., ans (Moore et al., 2008). Additionally, molecular 2009). Because plastid genomes are reduced with many investigations on some acquired genes in Cryptospori- genes related to photosynthesis being transferred to dium suggest that HGT may significantly affect the the algal nucleus, for these kleptoplasts to perform evolution of the biochemical system of apicomplexan photosynthesis, more proteins are needed than those pathogens (Striepen et al., 2002, 2004). encoded in their genomes. It has long been speculated

© 2012 Institute of Botany, Chinese Academy of Sciences 22 Journal of Systematics and Evolution Vol. 51 No. 1 2013 that these genes were transferred from the algal nucleus Much of the debate on the existence and the nature to sea slugs and their protein products are re‐imported of historical plastids centers on the controversial to kleptoplasts (Rumpho et al., 2000). Such algal assemblage “chromalveolates,” which includes both nuclear genes indeed have been identified in the sea plastid‐containing and plastid‐lacking taxa that are slug Elysia chlorotica (Pierce et al., 2007, 2012; presumably united by an ancestral algal endosymbiosis. Rumpho et al., 2008). In some other eukaryotes, The “chromalveolate” hypothesis has been under kleptoplasty can be achieved in a more complex constant debate and not been supported by more recent manner. For example, the ciliate Myrionecta rubra studies (Burki et al., 2008, 2012; Sanchez‐Puerta & preys on cryptophytes and retains their plastids. Delwiche, 2008; Bodyl et al., 2009; Parfrey et al., Myrionecta rubra, however, may also be consumed 2010). Because this hypothesis predicts plastid losses in by the dinoflagellate Dinophysis, which in turn steals plastid‐lacking members, algal genes in these taxa may the kleptoplasts. It has been shown that Dinophysis provide supporting evidence. Indeed, algal genes have acquired at least five genes from different sources been identified in multiple plastid‐lacking “chromal- to maintain the functionality of their kleptoplasts veolates” such as the apicomplexan Cryptosporidium (Wisecaver & Hackett, 2010). (Abrahamsen et al., 2004; Huang et al., 2004a), oomycetes (Tyler et al., 2006), and ciliates (Reyes‐ Prieto et al., 2008). However, whether all these genes 5 Algal genes and the historical distribution are derived from historical plastids remains an open of plastids question. For example, although strong cyanobacterial signal is found in oomycetes, statistical genomic Endosymbionts may be secondarily lost after their analyses show that such signal is not attributed to a genes were transferred to the nucleus. The spread of red algal endosymbiont as often postulated by the plastids in eukaryotes, either through primary or higher‐ “chromalveolate” hypothesis (Stiller et al., 2009). order endosymbioses, left algal and plastid genes (algal A related issue is whether algal genes in eukaryotes genes hereafter) as footprints in various taxa. Because can be derived from sources other than plastids or algal of the significant implication of plastid existence in endosymbionts. For instance, although algal genes in reconstructing eukaryotic phylogeny, tremendous ef- many plastid‐lacking eukaryotes are consistent with the forts have been made to infer historical plastids or algal scenario of historical plastids, they could actually have endosymbionts in some plastid‐lacking eukaryotes been acquired through feeding activities (Doolittle, (Huang et al., 2004a; Tyler et al., 2006; Maruyama 1998). An example is the choanoflagellate Monosiga et al., 2008, 2009; Reyes‐Prieto et al., 2008). These brevicollis, a unicellular protozoan organism that feeds efforts are often complicated by two major issues: (a) on bacteria and microscopic algae. Over 100 algal whether identified genes are indeed of algal origin and genes, mostly from haptophytes and diatoms, were (b) whether algal genes can be used as evidence for the identified in M. brevicollis (Nedelcu et al., 2008; Sun historical distribution of plastids. For example, it was et al., 2010; Sun & Huang, 2011). Another example is suggested that trypanosomatids, a group of eukaryotic the tunicate Ciona intestinalis, which also consumes parasites related to euglenoid algae, contained “plant‐ microscopic algae like M. brevicollis. Fourteen gene like” genes (Hannaert et al., 2003; Opperdoes & families in C. intestinalis are most likely of algal origin Michels, 2007) and thus possibly historical plastids and most of them appear to have been acquired by the (Hannaert et al., 2003). However, this suggestion has ancestral animal (Ni et al., 2012). Importantly, many not been supported by most later studies (Leander, of these acquired algal genes in M. brevicollis and 2004; Rogers & Keeling, 2004; Huang et al., 2004a; C. intestinalis ultimately are derived from bacteria, El‐Sayed et al., 2005; but also see Bodyl et al., 2010a). suggesting that HGT could be potentially frequent and The presence of plastid genes in several plastid‐lacking some beneficial genes may be transferred repeatedly eukaryotes also lent support to the hypothesis of a much under specific environments. Although the possibility more ancient primary plastid endosymbiosis in eukar- that algal genes in both species are derived from yotes and a super‐group “Plantae” that includes almost historical plastids cannot be entirely excluded, the fact all eukaryotes except opisthokonts (fungi and animals) that both Monosiga and Ciona prey on algae and and amoebozoans (Nozaki, 2005; Maruyama et al., bacteria renders gene acquisition through food sources 2008, 2009). Adding to this complex issue are some an at least equally possible scenario (Sun et al., 2010; other suggestions of historical plastids in fungi Yue & Huang, 2012). andanimals or in all eukaryotes (Yuan et al., 2008, The number of algal genes in M. brevicollis and 2010). C. intestinalis points to an issue about when algal genes

© 2012 Institute of Botany, Chinese Academy of Sciences HUANG & YUE: HGT in plants 23 may constitute valid evidence for historical plastids. In evolutionary novelties and, therefore, development and particular, M. brevicollis contains more algal genes than adaptation in different lineages of photosynthetic ciliates, which have similar habitats and feeding styles. eukaryotes. While algal genes in ciliates have been cited as support Plastids are the hallmark of photosynthetic for the “chromalveolate” hypothesis, no plastids have eukaryotes. HGT has played an important role in the been proposed for Monosiga or its close relatives under establishment of primary plastids and the evolution of any existing frameworks of eukaryotic evolution based other photosynthetic eukaryotes (Huang & Gogarten, on credible data. On the other hand, only a few algal 2008; Price et al., 2012). In an effort to investigate genes were found in the apicomplexan Cryptospori- ancient HGT in primary photosynthetic eukaryotes, 37 dium (Huang et al., 2004a). However, the scenario of genes acquired prior to the split of red algae and green plastid loss in Cryptosporidium was generally accepted plants were identified, including both novel genes and even before the finding of algal genes in them, largely those arising from homologous replacement (Huang & based on the fact that many other apicomplexans Gogarten, 2008). Importantly, the vast majority of these contain apicoplasts (Zhu et al., 2000). anciently acquired genes are related to the biogenesis At present, algal genes are mostly used as and functionality of plastids. Similar observations confirmatory evidence for historical plastids. For were also made in analyses of plastid proteomes example, genes of plastid origin have been found in (Archibald et al., 2003; Suzuki & Miyagishima, 2009). both Naegleria and fungi, the former being cited as The chlorarachniophyte Bigelowiella natans contains support for a more ancient origin of primary plastids, plastids of chlorophyte green algal origin, but many of whereas the latter considered as derived from inde- its plastid‐targeting proteins are derived from bacteria, pendent HGT events (Maruyama et al., 2009). Given red algae, or streptophyte green algae (Archibald that algal genes may exist in many plastid‐lacking et al., 2003). Similarly, over 100 plastid‐targeting eukaryotes, it is important to establish criteria to assess proteins common to both red algae and green plants the source of acquired genes so that historical plastids or were interpreted as acquired from prokaryotes, includ- endosymbionts can be reliably inferred. To this end, ing many from proteobacteria and chlamydiae (Suzuki Stiller (2011) proposed an approach that is based on & Miyagishima, 2009). These data indicate that gene rigorous statistical testing of a priori hypothesis against acquisition from miscellaneous sources might have a clearly defined control. This statistical approach contributed to the establishment of plastids and the makes great sense given that endosymbioses should optimization of plastid functionality in photosynthetic facilitate gene transfer and thus enhance phylogenetic eukaryotes (Huang & Gogarten, 2008). signal from endosymbionts in host nuclear genomes. Among all photosynthetic organisms, land plants However, it is unclear whether and how the enhanced are the most dominant on earth. One of the prominent signal from endosymbionts can be distinguished from features of land plants is their cell wall. Recent studies that arising from preferential feeding activities. Addi- also suggest that genes acquired from bacteria are tionally, because protein products of transferred genes involved in the formation of plant cell wall (Royo often function in original endosymbionts, loss of et al., 2000; Emiliani et al., 2009; Yin et al., 2011). plastids may potentially lead to loss of the originally Phenylalanine ammonia‐lyase (PAL) is a critical acquired algal genes. In such cases, it is also unclear enzyme of the general phenylpropanoid pathway in whether weaker algal signal will necessarily invalidate land plants that provides precursors for lignin monomer the possibility of historical plastids (Yue & Huang, biosynthesis. The land plant PAL was interpreted as 2012). resulting from HGT from soil bacteria to dikarya fungi, where it was secondarily routed to land plants (Emiliani et al., 2009). At least two genes acquired from 6 HGT in the origin and optimization of chlamydiae, including those encoding CMP‐KDO evolutionary novelties synthetase (CKS) and UDP‐D‐glucuronic acid 4‐ epimerase (UGlcAE), are involved in the biosynthesis Because the acquisition of foreign genes can of pectin (Royo et al., 2000; Yin et al., 2011). In potentially introduce novel metabolic capabilities or particular, UGlcAE converts UDP‐D‐glucuronic acid phenotypes, it is critical to assess the evolutionary into the activated form of galacturonic acid (GalA), the impact of HGT in recipient organisms. Thus far, most abundant sugar residue in pectin. It has been although our knowledge of HGT in photosynthetic estimated that GalA accounts for over 70% of total eukaryotes is still fragmental, available evidence pectin of the primary cell wall in some land plants suggests that HGT has contributed to the origin of (Mohnen, 2002). Therefore, it is apparent that these

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