Rapid Sequence Divergence Research Articles

The genome of Magnolia hypoleuca provides a new insight into cold tolerance and the evolutionary position of magnoliids.

Magnolia hypoleuca Sieb. & Zucc, a member of the Magnoliaceae of magnoliids, is one of the most economically valuable, phylogenetic and ornamental tree species in Eastern China. Here, the 1.64 Gb chromosome-level assembly covers 96.64% of the genome which is anchored to 19 chromosomes, with a contig N50 value of 1.71 Mb and 33,873 protein-coding genes was predicted. Phylogenetic analyses between M. hypoleuca and other 10 representative angiosperms suggested that magnoliids were placed as a sister group to the eudicots, rather than sister to monocots or both monocots and eudicots. In addition, the relative timing of the whole-genome duplication (WGD) events about 115.32 Mya for magnoliid plants. M. hypoleuca was found to have a common ancestor with M. officinalis approximately 23.4 MYA, and the climate change of OMT (Oligocene-Miocene transition) is the main reason for the divergence of M. hypoleuca and M. officinalis, which was along with the division of Japanese islands. Moreover, the TPS gene expansion observed in M. hypoleuca might contribute to the enhancement of flower fragrance. Tandem and proximal duplicates of younger age that have been preserved have experienced more rapid sequence divergence and a more clustered distribution on chromosomes contributing to fragrance accumulation, especially phenylpropanoid, monoterpenes and sesquiterpenes and cold tolerance. The stronger selective pressure drived the evolution of tandem and proximal duplicates toward plant self-defense and adaptation. The reference M. hypoleuca genome will provide insights into the evolutionary process of M. hypoleuca and the relationships between the magnoliids with monocots and eudicots, and enable us to delve into the fragrance and cold tolerance produced by M. hypoleuca and provide more robust and deep insight of how the Magnoliales evolved and diversified.

Divergence and introgression among the virilis group of Drosophila.

Speciation with gene flow is now widely regarded as common. However, the frequency of introgression between recently diverged species and the evolutionary consequences of gene flow are still poorly understood. The virilis group of Drosophila contains 12 species that are geographically widespread and show varying levels of prezygotic and postzygotic isolation. Here, we use de novo genome assemblies and whole-genome sequencing data to resolve phylogenetic relationships and describe patterns of introgression and divergence across the group. We suggest that the virilis group consists of three, rather than the traditional two, subgroups. Some genes undergoing rapid sequence divergence across the group were involved in chemical communication and desiccation tolerance, and may be related to the evolution of sexual isolation and adaptation. We found evidence of pervasive phylogenetic discordance caused by ancient introgression events between distant lineages within the group, and more recent gene flow between closely related species. When assessing patterns of genome-wide divergence in species pairs across the group, we found no consistent genomic evidence of a disproportionate role for the X chromosome as has been found in other systems. Our results show how ancient and recent introgressions confuse phylogenetic reconstruction, but may play an important role during early radiation of a group.

Adaptive sequence divergence forged new neurodevelopmental enhancers in humans

Searches for the genetic underpinnings of uniquely human traits have focused on human-specific divergence in conserved genomic regions, which reflects adaptive modifications of existing functional elements. However, the study of conserved regions excludes functional elements that descended from previously neutral regions. Here, we demonstrate that the fastest-evolved regions of the human genome, which we term "human ancestor quickly evolved regions" (HAQERs), rapidly diverged in an episodic burst of directional positive selection prior to the human-Neanderthal split, before transitioning to constraint within hominins. HAQERs are enriched for bivalent chromatin states, particularly in gastrointestinal and neurodevelopmental tissues, and genetic variants linked to neurodevelopmental disease. We developed a multiplex, single-cell invivo enhancer assay to discover that rapid sequence divergence in HAQERs generated hominin-unique enhancers in the developing cerebral cortex. We propose that a lack of pleiotropic constraints and elevated mutation rates poised HAQERs for rapid adaptation and subsequent susceptibility to disease.

Does rapid sequence divergence preclude RNA structure conservation in vertebrates?

Accelerated evolution of any portion of the genome is of significant interest, potentially signaling positive selection of phenotypic traits and adaptation. Accelerated evolution remains understudied for structured RNAs, despite the fact that an RNA’s structure is often key to its function. RNA structures are typically characterized by compensatory (structure-preserving) basepair changes that are unexpected given the underlying sequence variation, i.e., they have evolved through negative selection on structure. We address the question of how fast the primary sequence of an RNA can change through evolution while conserving its structure. Specifically, we consider predicted and known structures in vertebrate genomes. After careful control of false discovery rates, we obtain 13 de novo structures (and three known Rfam structures) that we predict to have rapidly evolving sequences—defined as structures where the primary sequences of human and mouse have diverged at least twice as fast (1.5 times for Rfam) as nearby neutrally evolving sequences. Two of the three known structures function in translation inhibition related to infection and immune response. We conclude that rapid sequence divergence does not preclude RNA structure conservation in vertebrates, although these events are relatively rare.

Phylogenetic convergence of phase separation and mitotic function in the disordered protein BuGZ.

Intrinsically disordered proteins (IDPs) effect biological function despite their sequence-encoded lack of preference for stable three-dimensional structure. Among their many functions, IDPs form membraneless cellular compartments through liquid-liquid phase separation (LLPS), also termed biomolecular condensation. The extent to which LLPS has been evolutionarily selected remains largely unknown, as the complexities of IDP evolution hamper progress. Unlike structured proteins, rapid sequence divergence typical of IDPs confounds inference of their biophysical or biological functions from comparative sequence analyses. Here, we leverage mitosis as a universal eukaryotic feature to interrogate condensate evolutionary history. We observe that evolution has conserved the ability for six homologs of the mitotic IDP BuGZ to undergo LLPS and to serve the same mitotic function, despite low sequence conservation. We also observe that cellular context may tune LLPS. The phylogenetic correlation of LLPS and mitotic function in one protein raises the possibility of an ancient evolutionary interplay between LLPS and biological function, dating back at least 1.6 billion years to the last common ancestor of plants and animals.

Rapid sequence evolution is associated with genetic incompatibilities in the plastid Clp complex.

Replacing the native clpP1 gene in the Nicotiana plastid genome with homologs from different donor species showed that the extent of genetic incompatibilities depended on the rate of sequence evolution. The plastid caseinolytic protease (Clp) complex plays essential roles in maintaining protein homeostasis and comprises both plastid-encoded and nuclear-encoded subunits. Despite the Clp complex being retained across green plants with highly conserved protein sequences in most species, examples of extremely accelerated amino acid substitution rates have been identified in numerous angiosperms. The causes of these accelerations have been the subject of extensive speculation but still remain unclear. To distinguish among prevailing hypotheses and begin to understand the functional consequences of rapid sequence divergence in Clp subunits, we used plastome transformation to replace the native clpP1 gene in tobacco (Nicotiana tabacum) with counterparts from another angiosperm genus (Silene) that exhibits a wide range in rates of Clp protein sequence evolution. We found that antibiotic-mediated selection could drive a transgenic clpP1 replacement from a slowly evolving donor species (S. latifolia) to homoplasmy but that clpP1 copies from Silene species with accelerated evolutionary rates remained heteroplasmic, meaning that they could not functionally replace the essential tobacco clpP1 gene. These results suggest that observed cases of rapid Clp sequence evolution are a source of epistatic incompatibilities that must be ameliorated by coevolutionary responses between plastid and nuclear subunits.

In silico identification of tetraspanins in monopisthocotylean (Platyhelminthes: Monogenea) parasites of fish.

Tetraspanins are a superfamily of transmembrane proteins that in flatworms have structural roles in the development, maturation or stability of the tegument. Several tetraspanins are considered as potential candidates for vaccines or drugs against helminths. Monopisthocotylean monogeneans are ectoparasites of fish that are health hazards for farmed fish. The aim of this study was to identify in silico putative tetraspanins in the genomic datasets of four monopisthocotylean species. The analysis predicted and classified 40 tetraspanins in Rhabdosynochus viridisi, 39 in Scutogyrus longicornis, 22 in Gyrodactylus salaris and 13 in Neobenedenia melleni, belonging to 13 orthologous groups. The high divergence of tetraspanins made it difficult to annotate their function. However, a conserved group was identified in different metazoan taxa. According to this study, metazoan tetraspanins can be divided into 17 monophyletic groups. Of the 114 monogenean tetraspanins, only seven were phylogenetically close to tetraspanins from non-platyhelminth metazoans, which suggests that this group of proteins shows rapid sequence divergence. The similarity of the monopisthocotylean tetraspanins was highest with trematodes, followed by cestodes and then free-living platyhelminths. In total, 27 monopisthocotylean-specific and 34 flatworm-specific tetraspanins were identified. Four monogenean tetraspanins were orthologous to TSP-1, which is a candidate for the development of vaccines and a potential pharmacological target in trematodes and cestodes. Although studies of tetraspanins in parasitic flatworms are scarce, this is an interesting group of proteins for the development of new methods to control monogeneans.

Propagation of a De Novo Gene under Natural Selection: Antifreeze Glycoprotein Genes and Their Evolutionary History in Codfishes.

The de novo birth of functional genes from non-coding DNA as an important contributor to new gene formation is increasingly supported by evidence from diverse eukaryotic lineages. However, many uncertainties remain, including how the incipient de novo genes would continue to evolve and the molecular mechanisms underlying their evolutionary trajectory. Here we address these questions by investigating evolutionary history of the de novo antifreeze glycoprotein (AFGP) gene and gene family in gadid (codfish) lineages. We examined AFGP phenotype on a phylogenetic framework encompassing a broad sampling of gadids from freezing and non-freezing habitats. In three select species representing different AFGP-bearing clades, we analyzed all AFGP gene family members and the broader scale AFGP genomic regions in detail. Codon usage analyses suggest that motif duplication produced the intragenic AFGP tripeptide coding repeats, and rapid sequence divergence post-duplication stabilized the recombination-prone long repetitive coding region. Genomic loci analyses support AFGP originated once from a single ancestral genomic origin, and shed light on how the de novo gene proliferated into a gene family. Results also show the processes of gene duplication and gene loss are distinctive in separate clades, and both genotype and phenotype are commensurate with differential local selective pressures.

Genome-wide identification, characterization and expression analysis of lineage-specific genes within Hanseniaspora yeasts.

Lineage-specific genes (LSGs) are defined as genes with sequences that are not significantly similar to those in any other lineage. LSGs have been proposed, and sometimes shown, to have significant effects in the evolution of biological function. In this study, two sets of Hanseniaspora spp. LSGs were identified by comparing the sequences of the Kloeckera apiculata genome and of 80 other yeast genomes. This study identified 344 Hanseniaspora-specific genes (HSGs) and 109 genes ('orphan genes') specific to K. apiculata. Three thousand three hundred thirty-oneK. apiculata genes that showed significant similarity to at least one sequence outside the Hanseniaspora were classified into evolutionarily conserved genes. We analyzed their sequence features, functional categories, gene origin, gene structure and gene expression. We also investigated the predicted cellular roles and Gene Ontology categories of the LSGs using functional inference. The patterns of the functions of LSGs do not deviate significantly from genome-wide average. The results showed that a few LSGs were formed by gene duplication, followed by rapid sequence divergence. Many of the HSGs and orphan genes exhibited altered expression in response to abiotic stress. Studying these LSGs might be helpful for understanding the molecular mechanism of yeast adaption.

Contrasting patterns of molecular evolution in metazoan germ line genes

BackgroundGerm lines are the cell lineages that give rise to the sperm and eggs in animals. The germ lines first arise from primordial germ cells (PGCs) during embryogenesis: these form from either a presumed derived mode of preformed germ plasm (inheritance) or from an ancestral mechanism of inductive cell-cell signalling (induction). Numerous genes involved in germ line specification and development have been identified and functionally studied. However, little is known about the molecular evolutionary dynamics of germ line genes in metazoan model systems.ResultsHere, we studied the molecular evolution of germ line genes within three metazoan model systems. These include the genus Drosophila (N=34 genes, inheritance), the fellow insect Apis (N=30, induction), and their more distant relative Caenorhabditis (N=23, inheritance). Using multiple species and established phylogenies in each genus, we report that germ line genes exhibited marked variation in the constraint on protein sequence divergence (dN/dS) and codon usage bias (CUB) within each genus. Importantly, we found that de novo lineage-specific inheritance (LSI) genes in Drosophila (osk, pgc) and in Caenorhabditis (pie-1, pgl-1), which are essential to germ plasm functions under the derived inheritance mode, displayed rapid protein sequence divergence relative to the other germ line genes within each respective genus. We show this may reflect the evolution of specialized germ plasm functions and/or low pleiotropy of LSI genes, features not shared with other germ line genes. In addition, we observed that the relative ranking of dN/dS and of CUB between genera were each more strongly correlated between Drosophila and Caenorhabditis, from different phyla, than between Drosophila and its insect relative Apis, suggesting taxonomic differences in how germ line genes have evolved.ConclusionsTaken together, the present results advance our understanding of the evolution of animal germ line genes within three well-known metazoan models. Further, the findings provide insights to the molecular evolution of germ line genes with respect to LSI status, pleiotropy, adaptive evolution as well as PGC-specification mode.

Mitonuclear coevolution as the genesis of speciation and the mitochondrial DNA barcode gap.

Mitochondrial genes are widely used in taxonomy and systematics because high mutation rates lead to rapid sequence divergence and because such changes have long been assumed to be neutral with respect to function. In particular, the nucleotide sequence of the mitochondrial gene cytochrome c oxidase subunit 1 has been established as a highly effective DNA barcode for diagnosing the species boundaries of animals. Rarely considered in discussions of mitochondrial evolution in the context of systematics, speciation, or DNA barcodes, however, is the genomic architecture of the eukaryotes: Mitochondrial and nuclear genes must function in tight coordination to produce the complexes of the electron transport chain and enable cellular respiration. Coadaptation of these interacting gene products is essential for organism function. I extend the hypothesis that mitonuclear interactions are integral to the process of speciation. To maintain mitonuclear coadaptation, nuclear genes, which code for proteins in mitochondria that cofunction with the products of mitochondrial genes, must coevolve with rapidly changing mitochondrial genes. Mitonuclear coevolution in isolated populations leads to speciation because population‐specific mitonuclear coadaptations create between‐population mitonuclear incompatibilities and hence barriers to gene flow between populations. In addition, selection for adaptive divergence of products of mitochondrial genes, particularly in response to climate or altitude, can lead to rapid fixation of novel mitochondrial genotypes between populations and consequently to disruption in gene flow between populations as the initiating step in animal speciation. By this model, the defining characteristic of a metazoan species is a coadapted mitonuclear genotype that is incompatible with the coadapted mitochondrial and nuclear genotype of any other population.

Loss of quaternary structure is associated with rapid sequence divergence in the OSBS family.

The rate of protein evolution is determined by a combination of selective pressure on protein function and biophysical constraints on protein folding and structure. Determining the relative contributions of these properties is an unsolved problem in molecular evolution with broad implications for protein engineering and function prediction. As a case study, we examined the structural divergence of the rapidly evolving o-succinylbenzoate synthase (OSBS) family, which catalyzes a step in menaquinone synthesis in diverse microorganisms and plants. On average, the OSBS family is much more divergent than other protein families from the same set of species, with the most divergent family members sharing <15% sequence identity. Comparing 11 representative structures revealed that loss of quaternary structure and large deletions or insertions are associated with the family's rapid evolution. Neither of these properties has been investigated in previous studies to identify factors that affect the rate of protein evolution. Intriguingly, one subfamily retained a multimeric quaternary structure and has small insertions and deletions compared with related enzymes that catalyze diverse reactions. Many proteins in this subfamily catalyze both OSBS and N-succinylamino acid racemization (NSAR). Retention of ancestral structural characteristics in the NSAR/OSBS subfamily suggests that the rate of protein evolution is not proportional to the capacity to evolve new protein functions. Instead, structural features that are conserved among proteins with diverse functions might contribute to the evolution of new functions.

Crosstalk and the evolution of specificity in two-component signaling

Two-component signaling (TCS) serves as the dominant signaling modality in bacteria. A typical pathway includes a sensor histidine kinase (HK) that phosphorylates a response regulator (RR), modulating its activity in response to an incoming signal. Most HKs are bifunctional, acting as both kinase and phosphatase for their substrates. Unlike eukaryotic signaling networks, there is very little crosstalk between bacterial TCS pathways; indeed, adding crosstalk to a pathway can have disastrous consequences for cell fitness. It is currently unclear exactly what feature of TCS necessitates this degree of pathway isolation. In this work we used mathematical models to show that, in the case of bifunctional HKs, adding a competing substrate to a TCS pathway will always reduce response of that pathway to incoming signals. We found that the pressure to maintain cognate signaling is sufficient to explain the experimentally observed "kinetic preference" of HKs for their cognate RRs. These findings imply a barrier to the evolution of new HK-RR pairs, because crosstalk is unavoidable immediately after the duplication of an existing pathway. We characterized a set of "near-neutral" evolutionary trajectories that minimize the impact of crosstalk on the function of the parental pathway. These trajectories predicted that crosstalk interactions should be removed before new input/output functionalities evolve. Analysis of HK sequences in bacterial genomes provided evidence that the selective pressures on the HK-RR interface are different from those experienced by the input domain immediately after duplication. This work thus provides a unifying explanation for the evolution of specificity in TCS networks.

Rapid Hepatitis C Virus Divergence among Chronically Infected Individuals

Here, we analyze the viral divergence among hepatitis C virus (HCV) chronic cases infected with genotype 1. The intrahost viral evolution was assessed by deep sequencing using the 454 Genome Sequencer platform. The results showed a rapid nucleotide sequence divergence. This notorious short-term viral evolution is of the utmost importance for the study of HCV transmission, because direct links between related samples were virtually lost. Thus, rapid divergence of HCV significantly affects genetic relatedness studies and outbreak investigations.

Long non‐coding RNAs coordinate cellular responses to stress

Following the initial discovery of the heat shock RNA omega (hsrω) gene of Drosophila melanogaster to be non-coding (nc) and also inducible by cell stress, other stress-inducible long non-coding RNAs (lncRNA) have been described in diverse organisms. In view of the rapid sequence divergence of lncRNAs, present knowledge of stress trasncriptome is limited and fragmented. Several known stress-related lncRNAs, associated with specific nuclear speckled domains or nucleolus, provide structural base for sequestering diverse RNA-processing/regulatory proteins. Others have roles in transcriptional or translational inhibition during stress or in signaling pathways; functions of several other lncRNAs are not yet known. Most stress-related lncRNAs act primarily by modulating activity of the proteins to which they bind or by sequestering specific sets of proteins away from the active pool. A common emerging theme is that a given lncRNA targets one or more protein/s with key role/s in the cascade of events triggered by the stress and therefore has a widespread integrative effect. Since proteins associate with RNA through short sequence motifs, the overall base sequence of functionally similar ncRNAs is often not conserved except for specific motifs. The rapid evolvability of ncRNA sequences provides elegant modules for adaptability to changing environment as binding of one or the other protein to ncRNA can alter its structure and functions in distinct ways. Thus the stress-related lncRNAs act as hubs in the cellular networks to coordinate activities of the members within and between different networks to maintain cellular homeostasis for survival or to trigger cell death.

A unique tRNA gene family and a novel, highly expressed ORF in the mitochondrial genome of the silver-lip pearl oyster, Pinctada maxima (Bivalvia: Pteriidae)

Characteristics of mitochondrial (mt) DNA such as gene content and arrangement, as well as mt tRNA secondary structure, are frequently used in comparative genomic analyses because they provide valuable phylogenetic information. However, most analyses do not characterize the relationship of tRNA genes from the same mt genome and, in some cases, analyses overlook possible novel open reading frames (ORFs) when the 13 expected protein-coding genes are already annotated. In this study, we describe the sequence and characterization of the complete mt genome of the silver-lip pearl oyster, Pinctada maxima. The 16,994-bp mt genome contains the same 13 protein-coding genes (PCGs) and two ribosomal RNA genes typical of metazoans. The gene arrangement, however, is completely distinct from that of all other available bivalve mt genomes, and a unique tRNA gene family is observed in this genome. The unique tRNA gene family includes two trnS−AGY and trnQ genes, a trnM isomerism, but it lacks trnS−CUN. We also report the first clear evidence of alloacceptor tRNA gene recruitment (trnP→trnS−AGY) in mollusks. In addition, a novel ORF (orfUR1) expressed at high levels is present in the mt genome of this pearl oyster. This gene contains a conserved domain, “Oxidored_q1_N”, which is a member of Complex I and thus may play an important role in key biological functions. Because orfUR1 has a very similar nucleotide composition and codon bias to that of other genes in this genome, we hypothesize that this gene may have been moved to the mt genome via gene transfer from the nuclear genome at an early stage of speciation of P. maxima, or it may have evolved as a result of gene duplication, followed by rapid sequence divergence. Lastly, a 319-bp region was identified as the possible control region (CR) even though it does not correspond to the longest non-coding region in the genome. Unlike other studies of mt genomes, this study compares the evolutionary patterns of all available bivalve mt tRNA and atp8 genes.

Ephemeral Association Between Gene CG5762 and Hybrid Male Sterility in Drosophila Sibling Species

Interspecies divergence in regulatory pathways may result in hybrid male sterility (HMS) when dominance and epistatic interactions between alleles that are functional within one genome are disrupted in hybrid genomes. The identification of genes contributing to HMS and other hybrid dysfunctions is essential for understanding the origin of new species (speciation). Previously, we identified a panel of male-specific loci misexpressed in sterile male hybrids of Drosophila simulans and D. mauritiana relative to parental species. In the current work, we attempt to dissect the genetic associations between HMS and one of the genes, CG5762, a Drosophila-unique locus characterized by rapid sequence divergence within the genus, presumably driven by positive natural selection. CG5762 is underexpressed in sterile backcross males compared with their fertile brothers. In CG5762 heterozygotes, the D. mauritiana allele is consistently overexpressed on both the D. simulans and D. mauritiana backcross genomic background, suggesting a cis-acting regulation factor. There is a significant association between heterozygosity and HMS in hybrid males from early but not later backcross generations. Microsatellite markers spanning CG5762 fail to associate with HMS. These observations lead to a conclusion that CG5762 is not a causative factor of HMS. Although genetic linkage between CG5762 and a neighboring causative introgression cannot be ruled out, it seems that the pattern is most consistent with CG5762 participating in epistatic interactions that are disrupted in flies with HMS.

Genomic survey of early responses to viruses in Atlantic salmon, Salmo salar L.

Viral diseases are one of the main problems and risk factors in aquaculture. At present diseases are diagnosed by detection of pathogens and clinical symptoms. Identification of genes involved in early responses to viruses is important for better knowledge of antiviral defence and development of diagnostic tools. The aim of this study was to search for gene markers common for viral infections in Atlantic salmon based on microarray analyses of a wide range of samples. Gene expression profiles from fish and cell cultures infected with different viruses and treated with the synthetic double-stranded RNA poly(I:C) were compared in order to identify virus responsive genes (VRG). The list of VRG defined in this study contained 117 genes with known or unidentified functions. Several genes, including the most highly ranked one (receptor transporting protein), had not been previously reported to be involved in antiviral defence. VRG were characterized by a rapid induction and low tissue specificity, and their expression levels were related to the viral load. Immunofluorescence analyses of proteins encoded by VRG in cardiac tissue of salmon with the viral disease cardiomyopathy syndrome (CMS) revealed a common expression pattern. In head kidney leukocytes VRG showed comparable or equal responses to CpG and poly(I:C), which mimic respectively bacterial DNA and viral RNA. Most VRG showed highly correlated expression with interferon-a (IFNa). Sequence comparison of salmon VRG with those from other species gave an understanding of the evolution of these genes, which showed a remarkably rapid sequence divergence in comparison with the entire proteome. VRG emerged both before and after separation of teleosts and tetrapods, and among genes found exclusively in fish species there were members of several multigene families: tripartite motif proteins, gig1- and gig2-like proteins. Several VRG, including genes with unknown functions and orthologs to mammalian RNA helicase RIG-I and chemokine C-X-C type 10, were present in cyprinid and salmonid fish but not in the phylogenetically advanced orders, suggesting that they have been lost in the evolution of Teleostei. Apparently, a number of genes involved in antiviral responses in salmon have acquired different functional roles in higher vertebrates.

Extreme genetic divergence is required for coreceptor switching in HIV-1 subtype C.

Coreceptor switching from CCR5 to CXCR4 is less common in subtype C HIV-1 infection than in subtype B for reasons that are unclear. We have examined sequential virus samples from a subtype C-infected child who had evidence of coreceptor switching. To examine HIV-1 envelope evolution towards CXCR4 usage, env sequences were correlated with phenotypic characteristics determined by entry assays, as well as the ability to use alternative coreceptors such as FPRL1, CCR3, CCR8 and others. The value of a phenotype predictor based on V3 sequences was also assessed. Ninety-three sequences revealed 3 distinct coexistent virus lineages and only some members of one lineage evolved to use CXCR4. These lineages also had diverse alternative coreceptor patterns including the ability to use FPRL1, CCR3, CCR8, APJ, CMKLR1, RDC-1, CXCR6, CCR1, GPCR1, GPR15 and CCR6. Coreceptor switching was associated with extensive and rapid sequence divergence in the V1/V2 region in addition to V3 changes. Furthermore, interlineage recombination within the C2 region resulted in low predictability of a V3 sequence-based phenotype algorithm, and highlighted the importance of V1/V2 and V3 sequences in coreceptor usage. These results suggest that the evolution to coreceptor switching in subtype C infection requires more mutations than other subtypes, and this contributes to the reduced incidence of R5X4 viruses.

Rapid Sequence and Expression Divergence Suggest Selection for Novel Function in Primate-Specific KRAB-ZNF Genes

Recent segmental duplications (SDs), arising from duplication events that occurred within the past 35-40 My, have provided a major resource for the evolution of proteins with primate-specific functions. KRAB zinc finger (KRAB-ZNF) transcription factor genes are overrepresented among genes contained within these recent human SDs. Here, we examine the structural and functional diversity of the 70 human KRAB-ZNF genes involved in the most recent primate SD events including genes that arose in the hominid lineage. Despite their recent advent, many parent-daughter KRAB-ZNF gene pairs display significant differences in zinc finger structure and sequence, expression, and splicing patterns, each of which could significantly alter the regulatory functions of the paralogous genes. Paralogs that emerged on the lineage to humans and chimpanzees have undergone more evolutionary changes per unit of time than genes already present in the common ancestor of rhesus macaques and great apes. Taken together, these data indicate that a substantial fraction of the recently evolved primate-specific KRAB-ZNF gene duplicates have acquired novel functions that may possibly define novel regulatory pathways and suggest an active ongoing selection for regulatory diversity in primates.