Molecular evolution of a primate-specific microRNA family

Lineage-specific microRNA (miRNA) families may contribute to developmental novelties during evolution. However, little is known about the origin and evolution of new miRNA families. We report evidence of an Alu-mediated rapid expansion of miRNA genes in a previously identified primate-specific miRNA family, drawn from sequencing and comparative analysis of 9 diverse primate species. Evolutionary analysis reveals similar divergence among miRNA copies whether they are within or between species, lineage-specific gain and loss of miRNAs, and gene pseudolization in multiple species. These observations support a birth-and-death process of miRNA genes in this family, implicating functional diversification during primate evolution. In addition, both secondary structure conservation and reduced single nucleotide polymorphisms density attest to functional constraint of this family in primates. Finally, we observed preferential expression of miRNAs in human placenta and fetal brain, suggesting a functional importance of this family for primate development.     

Differential Evolution of MAGE Genes Based on Expression Pattern and Selection Pressure

Starting from publicly-accessible datasets, we have utilized comparative and phylogenetic genome analyses to characterize the evolution of the human MAGE gene family. Our characterization of genomic structures in representative genomes of primates, rodents, carnivora, and macroscelidea indicates that both Type I and Type II MAGE genes have undergone lineage-specific evolution. The restricted expression pattern in germ cells of Type I MAGE orthologs is observed throughout evolutionary history. Unlike Type II MAGEs that have conserved promoter sequences, Type I MAGEs lack promoter conservation, suggesting that epigenetic regulation is a central mechanism for controlling their expression. Codon analysis shows that Type I but not Type II MAGE genes have been under positive selection. The combination of genomic and expression analysis suggests that Type 1 MAGE promoters and genes continue to evolve in the hominin lineage, perhaps towards functional diversification or acquiring additional specific functions, and that selection pressure at codon level is associated with expression spectrum.     

Glandular kallikreins of the cotton-top tamarin: molecular cloning of the gene encoding the tissue kallikrein

The glandular kallikrein family is composed of structurally related serine proteases. Studies show that the mouse family encompasses at least 14 highly conserved functional genes, but of these only the tissue kallikarein has a human ortholog. In man, the tissue kallikrein display high sequence similarity with prostate specific antigen and human glandular kallikrein 2, suggesting that they evolved after the separation of primates and rodents. A phylogenetic study of the genes encoding glandular kallikreins in species evolutionarily located between rodents and man may reveal interesting details on how the gene family evolved, which in turn could yield information about the function of the proteins. Therefore, we have initiated a study of the glandular kallikreins of the cotton-top tamarin (Saguinus oedipus), a New World Monkey. Here, we report the cloning and nucleotide sequence of one of these, the tissue kallikrein gene. The gene of 4.4 kb is composed of five exons, and the structure is 90% similar to that of the orthologous human gene. It gives rise to a polypeptide of 261 amino acids, including a signal peptide of 17 residues, a pro-piece of 7 residues, and the mature protein of 237 residues with an estimated molecular mass of 26.3 kD. The similarity to the human prostate specific antigen and human glandular kallikrein 2 genes is 73% and 72%, respectively, including introns and flanking regions. The lower similarity to these genes compared with the human tissue kallikrein gene indicates that they, or a progenitor to them, arose in primates prior to the separation of New and Old World monkeys. Genomic Southern blots also show that the cotton-top tamarin genome encompasses at least one more glandular kallikrein gene.     

Evolutionary Dynamics of the Interferon-Induced Transmembrane Gene Family in Vertebrates

Vertebrate interferon-induced transmembrane (IFITM) genes have been demonstrated to have extensive and diverse functions, playing important roles in the evolution of vertebrates. Despite observance of their functionality, the evolutionary dynamics of this gene family are complex and currently unknown. Here, we performed detailed evolutionary analyses to unravel the evolutionary history of the vertebrate IFITM family. A total of 174 IFITM orthologous genes and 112 pseudogenes were identified from 27 vertebrate genome sequences. The vertebrate IFITM family can be divided into immunity-related IFITM (IR-IFITM), IFITM5 and IFITM10 sub-families in phylogeny, implying origins from three different progenitors. In general, vertebrate IFITM genes are located in two loci, one containing the IFITM10 gene, and the other locus containing IFITM5 and various numbers of IR-IFITM genes. Conservation of evolutionary synteny was observed in these IFITM genes. Significant functional divergence was detected among the three IFITM sub-families. No gene duplication or positive selection was found in IFITM5 sub-family, implying the functional conservation of IFITM5 in vertebrate evolution, which is involved in bone formation. No IFITM5 locus was identified in the marmoset genome, suggesting a potential association with the tiny size of this monkey. The IFITM10 sub-family was divided into two groups: aquatic and terrestrial types. Functional divergence was detected between the two groups, and five IFITM10-like genes from frog were dispersed into the two groups. Both gene duplication and positive selection were observed in aquatic vertebrate IFITM10-like genes, indicating that IFITM10 might be associated with the adaptation to aquatic environments. A large number of lineage- and species-specific gene duplications were observed in IR-IFITM sub-family and positive selection was detected in IR-IFITM of primates and rodents. Because primates have experienced a long history of viral infection, such rapid expansion and positive selection suggests that the evolution of primate IR-IFITM genes is associated with broad-spectrum antiviral activity.     

Molecular Phylogeny,Evolution, and Functional Divergence of the LSD1-Like Gene Family: Inference from the Rice Genome

The identification of LSD1-like genes in parasite, green algae, moss, pine, and monocot and dicot species allowed us to trace the phylogenetic history of this gene family. Computational analysis showed that the diversification of members of this family could be dated back to the early stage of plant evolution. The evolution of plant LSD1-like genes was possibly shaped by two duplication events. These proteins, which contain three copies of the LSD1 zinc finger (zf-LSD1) domain within their entire polypeptides and play crucial roles in modulating disease defense and cell death, resulted from the second duplication. A gain of zf-LSD1 domain model was reasonable for explaining the origination of three-zf-LSD1 domain-containing proteins. The zf-LSD1 domain phylogeny showed that the middle (M) and C-terminal (C) domains originated from a common ancestor; the N-terminal (N) domain might be more ancient than the former two. The divergence of the N, M, and C domains was well before the monocot-dicot split. Coevolution analysis revealed that four intramolecular domain pairs, including the N domain and the interregion between the M and the C domains (INTER2), the M and C domain, the N- and C-terminus, and the M domain and C-terminus, possibly coevolved during the evolution of three-zf-LSD1 domain-containing proteins. The three zf-LSD1 domains are evolutionary conserved. Thus, the differences at the N- and C-terminus would be crucial for functional specificity of LSD1 genes. Strong functional constraints should work on the zf-LSD1 domains, whereas reduced functional constraint was found in the INTER2 region. Functional divergence analysis showed that three-zf-LSD1 domain-containing proteins were significantly functionally divergent from those proteins containing only one zf-LSD1 domain, a result demonstrating that shifted evolutionary rates between the two clusters were significantly different from each other. [Reviewing Editor: Dr. Joshua Plotkin]     

Evolutionary history and complementary selective relaxation of the duplicated PI genes in grasses

Gene duplication plays an important role in the evolution of organisms by allowing functional innovation and the divergence of duplicate genes. Previous studies found two PI-like genes in grass species, suggesting functional divergence between the paralogous copies. Here, we reconstructed the evolutionary history of two PI genes from major lineages of grasses and other monocot species, and demonstrated that two PI genes (PI1 and PI2) arose from a whole genome duplication that occurred in a common ancestor of extant grasses. Molecular evolutionary analyses at the family and tribal levels found strong purifying selection acting on two genes in grasses, consistent with the conserved class B function of the PI genes. Importantly, we detected different patterns of selective relaxation between the duplicated PI genes although no signature of positive selection was found. Likelihood ratio tests revealed that the ω ratio for M domain is significantly higher in PI1 than in PI2 but that for K domain is significantly higher in PI2 than in PI1. These findings imply that complementary selective relaxation occurs in two PI genes after duplication, and provide additional molecular evidence for the subfunctionalization of the duplicated PI genes in grasses.     

Molecular and evolutionary analyses of formyl peptide receptors suggest the absence of VNO-specific FPRs in primates

Formyl peptide receptors (FPRs) were observed to expand in rodents and were recently suggested as candidate vomeronasal chemo-sensory receptors. Since vomeronasal chemosensory receptors usually underwent positive selection and evolved concordantiy with the vomeronasal organ (VNO) morphology, we surveyed FPRs in primates in which VNO morphology is greatly diverse and thus it would provide us a clearer view of VNO-FPRs evolution. By screening available primate genome sequences, we obtained the FPR repertoires in representative primate species. As a result, we did not find FPR family size expansion in primates. Further analyses showed no evolutionary force variance between primates with or without VNO structure, which indicated that there was no functional divergence among primates FPRs. Our results suggest that primates lack the VNO-specific FPRs and the FPR expansion is not a common phenomenon in mammals outside rodent lineage, regardless of VNO complexity.     

The evolutionary conservation of the human chitotriosidase gene in rodents and primates

Chitinases have been identified in a variety of organisms ranging from prokaryotes to eukaryotes, known to specifically degrade chitin, an abundant polymer of N-acetylglucosamine. Recently a human chitinolytic enzyme called CHIT1 was discovered. CHIT1 is expressed by activated macrophages and hydrolyzes artificial chitotrioside substrates, but its specific function in humans is unknown, since it is generally believed that man completely lacks endogenous chitin and endogenous substrates for chitinases. An intriguing question is whether the chitotriosidase activity is just an evolutionary remnant or it has a physiological function in man. To test these hypotheses we utilized a "phylogenomic" approach performing accurate sequence analyses of this gene, coding for CHIT1, in rodents and primates. Inspecting the sequences available in public databases, we determined that this gene is conserved in rodents (mouse and rat) and primates (chimpanzee, gorilla, orangutan, gibbon, baboon, a common marmoset and black macaque). Moreover we found that a 24-base pair duplication that determines an enzymatically inactive human protein is not present in primates, suggesting that this polymorphism was created during human evolution. These results indicate that chitotriosidase is conserved across the evolutionary scale. Such conservation of the CHIT1 gene argues in favour of an important biological role.     

Molecular evolution of the primate developmental genes MSX1 and PAX9

In primates, the craniofacial skeleton and the dentition are marked by high levels of interspecific variation. Despite this, there are few comparative species studies conducted at the molecular level to investigate this functional diversity. We have determined nucleotide sequences of MSX1 and PAX9, two developmental genes, in a sample of 27 diverse primate species in order to identify coding or regulatory variation that may be associated with phenotypic diversity. Our analyses have identified four highly conserved noncoding sequences, including one that is conserved across primates and with dogs but not with mice. Although we find that substitution rates vary significantly across MSX1 exons, comparisons of nonsynonymous and synonymous substitution rates (dN/dS) suggest that, as a whole, MSX1 and PAX9 amino acid sequences have been under functional constraint throughout primate evolution. Compared to all other primates in our sample, our analysis of exon 1 in MSX1 finds an unusual pattern of amino acid substitution for Tarsius syrichta, a member of a lineage (tarsiers) that has many unique features among primates. For example, tarsiers are the only extant primates without deciduous incisors, and MSX1 is expressed exclusively in the incisor regions during the earliest stages of dental development. Our overall results provide insight into the utility of comparative species analyses of highly conserved developmental genes and their roles in the evolution of complex phenotypes.     

Natural history and functional divergence of protein tyrosine kinases

Cellular signaling is important for many biological processes including growth, differentiation, adhesion, motility and apoptosis. The protein tyrosine kinase (PTK) supergene family is the key mediator in cellular signaling in metazoans, directly associated with a variety of human diseases. All PTKs contain a highly conserved catalytic kinase domain, in spite of variable multi-domain structures. Within each PTK gene family, members exhibit functional divergence in substrate-specificity or temporal/tissue-specific expression, although their primary function is conserved. After conducting phylogenetic analysis on major PTK gene families, we found that the expanding of each PTK family was likely caused by gene or genome duplication event(s) that occurred before the emergence of teleosts but after the vertebrate-amphioxus split. We further investigated the evolutionary pattern of functional divergence after gene duplication in those gene families. Our results show that site-specific shifted evolutionary rate (altered functional constraint) is a common pattern in PTK gene family evolution.     

Natural selection on functional modules, a genome-wide analysis

Classically, the functional consequences of natural selection over genomes have been analyzed as the compound effects of individual genes. The current paradigm for large-scale analysis of adaptation is based on the observed significant deviations of rates of individual genes from neutral evolutionary expectation. This approach, which assumed independence among genes, has not been able to identify biological functions significantly enriched in positively selected genes in individual species. Alternatively, pooling related species has enhanced the search for signatures of selection. However, grouping signatures does not allow testing for adaptive differences between species. Here we introduce the Gene-Set Selection Analysis (GSSA), a new genome-wide approach to test for evidences of natural selection on functional modules. GSSA is able to detect lineage specific evolutionary rate changes in a notable number of functional modules. For example, in nine mammal and Drosophilae genomes GSSA identifies hundreds of functional modules with significant associations to high and low rates of evolution. Many of the detected functional modules with high evolutionary rates have been previously identified as biological functions under positive selection. Notably, GSSA identifies conserved functional modules with many positively selected genes, which questions whether they are exclusively selected for fitting genomes to environmental changes. Our results agree with previous studies suggesting that adaptation requires positive selection, but not every mutation under positive selection contributes to the adaptive dynamical process of the evolution of species.     

Various aspects of evolution of Alu repeats in mammals

A complex study on various evolutionary peculiarities of the mammalia dispersed Alu repeats (Alu repeats of primates and B1 of rodents) has been carried out by phylogenetic analysis. A phylogenetic tree, containing the 7SL RNA genes and the Alu repeats of primates and rodents has been constructed. It has been shown that the branch of the phyletic line leading to the Alu repeats of primates and B1 of rodents from the 7SL RNA genes occurred after the divergence of the 7SL RNA genes of amphibia and mammalia, but before the divergence of the 7SL RNA genes of primates and rodents (250.10 years ago). A statistically reliable slowing down in the evolutionary rates of one of two monomers for the human Alu repeats has been proved. It may be caused by the functional load of the corresponding monomer in connection with the presence of a definit regulatory site in it.     

Molecular evolution of <Emphasis Type="Italic">PKD2</Emphasis> gene family in mammals

PKD2 gene encodes a critical cation channel protein that plays important roles in various developmental processes and is usually evolutionarily conserved. In the present study, we analyzed the evolutionary patterns of PKD2 and its homologous genes (PKD2L1, PKD2L2) from nine mammalian species. In this study, we demonstrated the orthologs of PKD2 gene family evolved under a dominant purifying selection force. Our results in combination with the reported evidences from functional researches suggested the entire PKD2 gene family are conserved and perform essential biological roles during mammalian evolution. In rodents, PKD2 gene family members appeared to have evolved more rapidly than other mammalian lineages, probably resulting from relaxation of purifying selection. However, positive selection imposed on synonymous sites also potentially contributed to this case. For the paralogs, our results implied that PKD2L2 genes evolved under a weaker purifying selection constraint than PKD2 and PKD2L1 genes. Interestingly, some loop regions of transmembrane domain of PKD2L2 exhibited higher P _N/P _S ratios than expected, suggesting these regions are more functional divergent in organisms and worthy of special attention. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Chun Ye, Huan Sun have contributed equally to this work.