Higher Rates Of Nonsynonymous Substitution Research Articles

Differentiated evolutionary rates in alternative exons and the implications for splicing regulation.

BackgroundAlternatively spliced exons play an important role in the diversification of gene function in most metazoans and are highly regulated by conserved motifs in exons and introns. Two contradicting properties have been associated to evolutionary conserved alternative exons: higher sequence conservation and higher rate of non-synonymous substitutions, relative to constitutive exons. In order to clarify this issue, we have performed an analysis of the evolution of alternative and constitutive exons, using a large set of protein coding exons conserved between human and mouse and taking into account the conservation of the transcript exonic structure. Further, we have also defined a measure of the variation of the arrangement of exonic splicing enhancers (ESE-conservation score) to study the evolution of splicing regulatory sequences. We have used this measure to correlate the changes in the arrangement of ESEs with the divergence of exon and intron sequences.ResultsWe find evidence for a relation between the lack of conservation of the exonic structure and the weakening of the sequence evolutionary constraints in alternative and constitutive exons. Exons in transcripts with non-conserved exonic structures have higher synonymous (dS) and non-synonymous (dN) substitution rates than exons in conserved structures. Moreover, alternative exons in transcripts with non-conserved exonic structure are the least constrained in sequence evolution, and at high EST-inclusion levels they are found to be very similar to constitutive exons, whereas alternative exons in transcripts with conserved exonic structure have a dS significantly lower than average at all EST-inclusion levels. We also find higher conservation in the arrangement of ESEs in constitutive exons compared to alternative ones. Additionally, the sequence conservation at flanking introns remains constant for constitutive exons at all ESE-conservation values, but increases for alternative exons at high ESE-conservation values.ConclusionWe conclude that most of the differences in dN observed between alternative and constitutive exons can be explained by the conservation of the transcript exonic structure. Low dS values are more characteristic of alternative exons with conserved exonic structure, but not of those with non-conserved exonic structure. Additionally, constitutive exons are characterized by a higher conservation in the arrangement of ESEs, and alternative exons with an ESE-conservation similar to that of constitutive exons are characterized by a conservation of the flanking intron sequences higher than average, indicating the presence of more intronic regulatory signals.

Relaxation of Functional Constraint on Light-Independent Protochlorophyllide Oxidoreductase in Thuja

The light-independent protochlorophyllide oxidoreductase (DPOR) plays a key role in the ability of nonflowering plants and algae to synthesize chlorophyll in darkness. This enzyme consists of three subunits encoded by the chlB, chlL, and chlN genes in the plastid genome. Previously, we found a high nonsynonymous substitution rate (dN) of the chlL gene in the lineage of Thuja standishii, a conifer belonging to the Cupressaceae. Here we revealed that the acceleration of dN in the chlL occurred as well in other species of Thuja, Thuja occidentalis and Thuja plicata. In addition, dark-grown seedlings of T. occidentalis were found to exhibit a pale yellowish color, and their chlorophyll concentration was much lower than that of other species of Cupressaceae. The results suggested that the species of Thuja have lost the ability to synthesize chlorophyll in darkness, and the functional constraint on the DPOR would thus be expected to be relaxed in this genus. Therefore, we expected to find that the evolutionary rates of all subunits of DPOR would in this case be accelerated. Sequence analyses of the chlN and chlB (encoding the other subunits of DPOR) in 18 species of Cupressaceae revealed that the dN of the chlN gene was accelerated in Thuja as was the dN of the chlL gene, but the dN of the chlB gene did not appear to differ significantly among the species of Cupressaceae. Sequencing of reverse transcription-polymerase chain reaction (RT-PCR) products of these genes showed that RNA editing was rare and unlikely to have contributed to the acceleration. Moreover, the RT-PCR analysis indicated that all chl genes were still transcriptionally active in T. occidentalis. Based on these results, it appears that species of Thuja still bear the DPOR protein, although the enzyme has lost its activity because of nonsynonymous mutations of some of the chl genes. The lack of acceleration of the dN of the chlB gene might be accounted for by various unknown functions of its gene product.

Parallels and contrasts between symbiotic bacteria and bacterial-derived organelles: evidence from Buchnera, the bacterial symbiont of aphids

Bacteria of the genus Buchnera (γ-Proteobacteria, allied with Escherichia coli) occur exclusively in symbiosis with aphids. Buchnera is advantageous to aphids because it over-produces essential amino acids, which are made available to the insect tissues. The congruence of the phylogenies of Buchnera and aphids, together with dating from the aphid fossil record, suggests that Buchnera has been transmitted vertically down the aphid generations for 160–280 million years. Despite this long history in symbiosis, the Buchnera genome does not exhibit the reduction and simplification characteristic of bacteria-derived organelles. The principal genetic reorganisation, relative to E. coli, relates to amino acid over-production. Buchnera in advanced aphid families has genes for enzymes in the synthesis of two essential amino acids, tryptophan and leucine, amplified on plasmids. The Buchnera genome is also characterized by high AT content and high rate of nonsynonymous substitutions in protein-coding genes. The latter may include deleterious mutations that accumulate because (as a consequence of vertical transmission) Buchnera lacks the opportunity for recombination.

Comparative analysis of chimpanzee and human Y chromosomes unveils complex evolutionary pathway

The mammalian Y chromosome has unique characteristics compared with the autosomes or X chromosomes. Here we report the finished sequence of the chimpanzee Y chromosome (PTRY), including 271 kb of the Y-specific pseudoautosomal region 1 and 12.7 Mb of the male-specific region of the Y chromosome. Greater sequence divergence between the human Y chromosome (HSAY) and PTRY (1.78%) than between their respective whole genomes (1.23%) confirmed the accelerated evolutionary rate of the Y chromosome. Each of the 19 PTRY protein-coding genes analyzed had at least one nonsynonymous substitution, and 11 genes had higher nonsynonymous substitution rates than synonymous ones, suggesting relaxation of selective constraint, positive selection or both. We also identified lineage-specific changes, including deletion of a 200-kb fragment from the pericentromeric region of HSAY, expansion of young Alu families in HSAY and accumulation of young L1 elements and long terminal repeat retrotransposons in PTRY. Reconstruction of the common ancestral Y chromosome reflects the dynamic changes in our genomes in the 5-6 million years since speciation.

Conservation of pregnancy-specific glycoprotein (PSG) N domains following independent expansions of the gene families in rodents and primates

BackgroundRodent and primate pregnancy-specific glycoprotein (PSG) gene families have expanded independently from a common ancestor and are expressed virtually exclusively in placental trophoblasts. However, within each species, it is unknown whether multiple paralogs have been selected for diversification of function, or for increased dosage of monofunctional PSG. We analysed the evolution of the mouse PSG sequences, and compared them to rat, human and baboon PSGs to attempt to understand the evolution of this complex gene family.ResultsPhylogenetic tree analyses indicate that the primate N domains and the rodent N1 domains exhibit a higher degree of conservation than that observed in a comparison of the mouse N1 and N2 domains, or mouse N1 and N3 domains. Compared to human and baboon PSG N domain exons, mouse and rat PSG N domain exons have undergone less sequence homogenisation. The high non-synonymous substitution rates observed in the CFG face of the mouse N1 domain, within a context of overall conservation, suggests divergence of function of mouse PSGs. The rat PSG family appears to have undergone less expansion than the mouse, exhibits lower divergence rates and increased sequence homogenisation in the CFG face of the N1 domain. In contrast to most primate PSG N domains, rodent PSG N1 domains do not contain an RGD tri-peptide motif, but do contain RGD-like sequences, which are not conserved in rodent N2 and N3 domains.ConclusionRelative conservation of primate N domains and rodent N1 domains suggests that, despite independent gene family expansions and structural diversification, mouse and human PSGs retain conserved functions. Human PSG gene family expansion and homogenisation suggests that evolution occurred in a concerted manner that maintains similar functions of PSGs, whilst increasing gene dosage of the family as a whole. In the mouse, gene family expansion, coupled with local diversification of the CFG face, suggests selection both for increased gene dosage and diversification of function. Partial conservation of RGD and RGD-like tri-peptides in primate and rodent N and N1 domains, respectively, supports a role for these motifs in PSG function.

Molecular Evolution of Duplicated Ray Finned Fish HoxA Clusters: Increased Synonymous Substitution Rate and Asymmetrical Co-divergence of Coding and Non-coding Sequences

In this study the molecular evolution of duplicated HoxA genes in zebrafish and fugu has been investigated. All 18 duplicated HoxA genes studied have a higher non-synonymous substitution rate than the corresponding genes in either bichir or paddlefish, where these genes are not duplicated. The higher rate of evolution is not due solely to a higher non-synonymous-to-synonymous rate ratio but to an increase in both the non-synonymous as well as the synonymous substitution rate. The synonymous rate increase can be explained by a change in base composition, codon usage, or mutation rate. We found no changes in nucleotide composition or codon bias. Thus, we suggest that the HoxA genes may experience an increased mutation rate following cluster duplication. In the non-Hox nuclear gene RAG1 only an increase in non-synonymous substitutions could be detected, suggesting that the increased mutation rate is specific to duplicated Hox clusters and might be related to the structural instability of Hox clusters following duplication. The divergence among paralog genes tends to be asymmetric, with one paralog diverging faster than the other. In fugu, all b-paralogs diverge faster than the a-paralogs, while in zebrafish Hoxa-13a diverges faster. This asymmetry corresponds to the asymmetry in the divergence rate of conserved non-coding sequences, i.e., putative cis-regulatory elements. These results suggest that the 5' HoxA genes in the same cluster belong to a co-evolutionary unit in which genes have a tendency to diverge together.

MHC Class II DRB Variability and Parasite Load in the Striped Mouse (Rhabdomys pumilio) in the Southern Kalahari

Major histocompatibility complex (MHC) variability is believed to be maintained by pathogen-driven selection, mediated either through heterozygous advantage or frequency-dependent selection. However, empirical support for these hypotheses under natural conditions is rare. In this study, we investigated the genetic constitution of the functionally important MHC class II gene (DRB exon 2) and the parasite load in a population of the striped mouse (Rhabdomys pumilio) in the Southern Kalahari. Fifty-eight individuals were genetically examined and the endoparasite load was quantified by counting fecal helminth eggs by using a modified McMaster technique. Thirty-four animals (58.6%) were infected. We identified 20 different MHC alleles with high levels of sequence divergence between alleles. Particularly, the antigen-binding sites revealed a significant higher rate of nonsynonymous substitutions (d(N)) than synonymous substitutions (d(S)), giving strong evidence of balancing selection. Heterozygosity did influence the infection status (being infected or not) and the individual fecal egg count (FEC) value with significantly higher values observed in homozygous individuals. Furthermore, a positive relationship was found between specific alleles and parasite load. The allele Rhpu-DRB*1 significantly occurred more frequently in infected individuals and in individuals with high FEC values (high parasite load). Individuals with the allele Rhpu-DRB*1 had a 1.5-fold higher chance of being infected than individuals without this allele (odds ratio test, P < 0.05). Contrarily, the allele Rhpu-DRB*8 significantly occurred more frequent in individuals with low FEC values. Our results support the hypotheses that MHC polymorphism in R. pumilio is maintained through pathogen-driven selection acting by both heterozygosity advantage and frequency-dependent selection.

Phylogeny and intraspecific variability of holoparasitic Orobanche (Orobanchaceae) inferred from plastid rbcL sequences

The rbcL sequences of 106 specimens representing 28 species of the four recognized sections of Orobanche were analyzed and compared. Most sequences represent pseudogenes with premature stop codons. This study confirms that the American lineage (sects. Gymnocaulis and Myzorrhiza) contains potentially functional rbcL-copies with intact open reading frames and low rates of non-synonymous substitutions. For the first time, this is also shown for a member of the Eurasian lineage, O. coerulescens of sect. Orobanche, while all other investigated species of sects. Orobanche and Trionychon contain pseudogenes with distorted reading frames and significantly higher rates of non-synonymous substitutions. Phylogenetic analyses of the rbcL sequences give equivocal results concerning the monophyly of Orobanche, and the American lineage might be more closely related to Boschniakia and Cistanche than to the other sections of Orobanche. Additionally, species of sect. Trionychon phylogenetically nest in sect. Orobanche. This is in concordance with results from other plastid markers ( rps2 and matK), but in disagreement with other molecular (nuclear ITS), morphological, and karyological data. This might indicate that the ancestor of sect. Trionychon has captured the plastid genome, or parts of it, of a member of sect. Orobanche. Apart from the phylogenetically problematic position of sect. Trionychon, the phylogenetic relationships within sect. Orobanche are similar to those inferred from nuclear ITS data and are close to the traditional groupings traditionally recognized based on morphology. The intraspecific variation of rbcL is low and is neither correlated with intraspecific morphological variability nor with host range. Ancestral character reconstruction using parsimony suggests that the ancestor of O. sect. Orobanche had a narrow host range.

Comparative analysis of the receptor-like kinase family in Arabidopsis and rice.

Receptor-like kinases (RLKs) belong to the large RLK/Pelle gene family, and it is known that the Arabidopsis thaliana genome contains >600 such members, which play important roles in plant growth, development, and defense responses. Surprisingly, we found that rice (Oryza sativa) has nearly twice as many RLK/Pelle members as Arabidopsis does, and it is not simply a consequence of a larger predicted gene number in rice. From the inferred phylogeny of all Arabidopsis and rice RLK/Pelle members, we estimated that the common ancestor of Arabidopsis and rice had >440 RLK/Pelles and that large-scale expansions of certain RLK/Pelle members and fusions of novel domains have occurred in both the Arabidopsis and rice lineages since their divergence. In addition, the extracellular domains have higher nonsynonymous substitution rates than the intracellular domains, consistent with the role of extracellular domains in sensing diverse signals. The lineage-specific expansions in Arabidopsis can be attributed to both tandem and large-scale duplications, whereas tandem duplication seems to be the major mechanism for recent expansions in rice. Interestingly, although the RLKs that are involved in development seem to have rarely been duplicated after the Arabidopsis-rice split, those that are involved in defense/disease resistance apparently have undergone many duplication events. These findings led us to hypothesize that most of the recent expansions of the RLK/Pelle family have involved defense/resistance-related genes.

USING COMPARATIVE GENOMIC DATA TO TEST FOR FAST-X EVOLUTION

Abstract Genes may acquire nonsynonymous substitutions more rapidly when X-linked than when autosomal, but evidence for “fast-X evolution” has been elusive. Fast-X evolution could explain the disproportionate contribution of X-linked genes to hybrid sterility and other traits. Here, we use a comparative genomic approach, with sequences of 30--110 genes in four Drosophila species, to test for fast-X evolution. Specifically, the 3L autosome arm in D. melanogaster and D. simulans is homologous to the right arm of the X chromosome in D. pseudoobscura and D. miranda. We executed two paired comparisons to determine how often genes on this chromosome arm exhibit higher rates of nonsynonymous substitution in the D. pseudoobscura species group, as predicted by fast-X evolution. We found a statistically significant pattern consistent with fast-X evolution in one comparison and a similar trend in the other comparison. Variation in functional constraints across genes may have masked the signature of fast-X evolution in some previous studies, and we conclude paired comparisons are more powerful for examining rates of evolution of genes when X-linked over autosomal.

USING COMPARATIVE GENOMIC DATA TO TEST FOR FAST-X EVOLUTION

Genes may acquire nonsynonymous substitutions more rapidly when X-linked than when autosomal, but evidence for "fast-X evolution" has been elusive. Fast-X evolution could explain the disproportionate contribution of X-linked genes to hybrid sterility and other traits. Here, we use a comparative genomic approach, with sequences of 30-110 genes in four Drosophila species, to test for fast-X evolution. Specifically, the 3L autosome arm in D. melanogaster and D. simulans is homologous to the right arm of the X chromosome in D. pseudoobscura and D. miranda. We executed two paired comparisons to determine how often genes on this chromosome arm exhibit higher rates of nonsynonymous substitution in the D. pseudoobscura species group, as predicted by fast-X evolution. We found a statistically significant pattern consistent with fast-X evolution in one comparison and a similar trend in the other comparison. Variation in functional constraints across genes may have masked the signature of fast-X evolution in some previous studies, and we conclude paired comparisons are more powerful for examining rates of evolution of genes when X-linked over autosomal.

Rapidly evolving genes in human. I. The glycophorins and their possible role in evading malaria parasites.

In an attempt to identify all fast-evolving genes between human and other primates, we found three glycophorins, GPA, GPB, and GPE, to have the highest rate of nonsynonymous substitutions among the 280 genes surveyed. The Ka/Ks ratios are generally greater than 3 for GPA, GPB, and GPE in human, chimpanzee, and gorilla, indicating positive selection. The uniformly high substitution rate across loci can be explained by the frequent sequence exchanges among genes. GPA is the receptor for the binding ligand EBA-175 of the malaria parasite, Plasmodium falciparum. The levels of nonsynonymous divergence and polymorphism of EBA-175 are also the highest in the genome of P. falciparum. We hypothesize that GPA has been evolving rapidly to evade malaria parasites. Both the high rate of nonsynonymous substitutions and the frequent interlocus conversions may be means of evasion. The support for the evasion hypothesis is still indirect, but, unlike other hypotheses, it can be tested specifically and systematically.

Patterns of polymorphism in the MHC class II of a non-passerine bird, the great snipe (Gallinago media).

The genomic organisation of the major histocompatibility complex (MHC) seems to vary considerably between different bird species. In order to understand this variation it is important to gather information from different species. We have, for the first time, investigated MHC class II polymorphism in a wader species, the great snipe (Gallinago media). Eleven alleles were found in five sequenced individuals; these come from at least three different loci, but RFLP data suggest that a larger number of genes may be present. For MHC genes, amino acid substitutions followed the, for MHC genes, general pattern of high non-synonymous substitution rates in peptide-binding regions, suggesting that the sequenced alleles may be expressed. The number of genes, lengths of introns and exon sequences of the great snipe MHC seem to be intermediate between those of chicken and passerine birds.

Substitution rates in the X- and Y-linked genes of the plants, Silene latifolia and S. dioica.

Theory predicts that selection should be less effective in the nonrecombining genes of Y-chromosomes, relative to the situation for genes on the other chromosomes, and this should lead to the accumulation of deleterious nonsynonymous substitutions. In addition, synonymous substitution rates may differ between X- and Y-linked genes because of the male-driven evolution effect and also because of actual differences in per-replication mutation rates between the sex chromosomes. Here, we report the first study of synonymous and nonsynonymous substitution rates on plant sex chromosomes. We sequenced two pairs of sex-linked genes, SlX1-SlY1 and SlX4-SlY4, from dioecious Silene latifolia and S. dioica, and their non-sex-linked homologues from nondioecious S. vulgaris and Lychnis flos-jovis, respectively. The rate of nonsynonymous substitutions in the SlY4 gene is significantly higher than that in the SlX4 gene. Silent substitution rates are also significantly higher in both Y-linked genes, compared with their X-linked homologues. The higher nonsynonymous substitution rate in the SlY4 gene is therefore likely to be caused by a mutation rate difference between the sex chromosomes. The difference in silent substitution rates between the SlX4 and SlY4 genes is too great to be explained solely by a higher per-generation mutation rate in males than females. It is thus probably caused by a difference in per-replication mutation rates between the sex chromosomes. This suggests that the local mutation rate can change in a relatively short evolutionary time.

Origin and evolution of viral interleukin-10 and other DNA virus genes with vertebrate homologues.

Phylogenies of gene families including members in both vertebrates and DNA viruses of the poxvirus and/or herpesvirus families showed that the viral genes originated at widely different times over the history of life. Certain of these viral genes (for example, the genes encoding the large and small subunits of ribonucleoside-diphosphate reductase) originated before animals diverged from fungi, while others originated much more recently. The most striking examples of recent origin involved viral genes encoding the cytokine interleukin-10 (IL-10), which originated independently in viruses at least three times since the divergence of the orders of eutherian mammals, presumably by viral capture of host genes. In certain domains, viral IL-10 genes showed significantly higher rates of nonsynonymous substitution than their nearest mammalian homologues. Though the mutation rate in these viral genes is up to 20 times that of the corresponding mammalian genes, a high mutation rate alone did not account for these differences because they were not seen in all domains. Rather, in certain domains it appears that functional constraints present in the case of mammalian IL-10 are relaxed in the case of the viral homologues. Furthermore, a nonrandom pattern of change with respect to amino acid residue charge in the N-terminal portion of the mature protein has occurred repeatedly in independently derived viral IL-10 genes, strongly suggesting that positive selection has led to divergence of this functionally important domain in viral IL-10.

Molecular evolution of the teosinte branched gene among maize and related grasses.

Several authors have proposed that changes in a small number of regulatory genes may be sufficient for the evolution of novel morphologies. Recent analyses have indicated that teosinte branched1 (tb1), a putative bHLH transcription factor, played such a role during the morphological evolution of maize from its wild ancestor, teosinte. To address whether or not tb1 played a similar role during the evolution of the Andropogoneae, the tribe to which maize belongs, and to examine the rate and pattern of tb1 evolution within this tribe, we analyzed tb1-like sequences from 23 members of the Andropogoneae and five other grasses. Our analysis revealed that the TB1 protein evolves slowly within three conserved domains but rapidly outside these domains. The nonconserved regions of the gene are characterized by both a high nonsynonymous substitution rate and frequent indels. The ratio of nonsynonymous substitutions per nonsynonymous site (d(N)) to synonymous substitutions per synonymous site (d(S)) was not significantly greater than 1.0, providing no evidence for positive selection. However, the d(N)/d(S) ratio varied significantly among lineages and was high compared with those of other plant nuclear genes. Variation in the d(N)/d(S) ratio among the Andropogoneae could be explained by unequal levels of purifying selection among lineages. Consistent with this interpretation, the rate of nonsynonymous substitution differed along several lineages, while the synonymous substitution rate did not differ significantly. Finally, using tb1, we examined phylogenetic relationships within the Andropogoneae. The phylogeny suggests that the tribe underwent a rapid radiation during its early history and that the monoecious Andropogoneae are polyphyletic.

Dichotomy of single-nucleotide polymorphism haplotypes in olfactory receptor genes and pseudogenes.

Substantial efforts are focused on identifying single-nucleotide polymorphisms (SNPs) throughout the human genome, particularly in coding regions (cSNPs), for both linkage disequilibrium and association studies. Less attention, however, has been directed to the clarification of evolutionary processes that are responsible for the variability in nucleotide diversity among different regions of the genome. We report here the population sequence diversity of genomic segments within a 450-kb cluster of olfactory receptor (OR) genes on human chromosome 17. We found a dichotomy in the pattern of nucleotide diversity between OR pseudogenes and introns on the one hand and the closely interspersed intact genes on the other. We suggest that weak positive selection is responsible for the observed patterns of genetic variation. This is inferred from a lower ratio of polymorphism to divergence in genes compared with pseudogenes or introns, high non-synonymous substitution rates in OR genes, and a small but significant overall reduction in variability in the entire OR gene cluster compared with other genomic regions. The dichotomy among functionally different segments within a short genomic distance requires high recombination rates within this OR cluster. Our work demonstrates the impact of weak positive selection on human nucleotide diversity, and has implications for the evolution of the olfactory repertoire.

Positive selection and propeptide repeats promote rapid interspecific divergence of a gastropod sperm protein.

Male-specific proteins have increasingly been reported as targets of positive selection and are of special interest because of the role they may play in the evolution of reproductive isolation. We report the rapid interspecific divergence of cDNA encoding a major acrosomal protein of unknown function (TMAP) of sperm from five species of teguline gastropods. A mitochondrial DNA clock (calibrated by congeneric species divided by the Isthmus of Panama) estimates that these five species diverged 2-10 MYA. Inferred amino acid sequences reveal a propeptide that has diverged rapidly between species. The mature protein has diverged faster still due to high nonsynonymous substitution rates (> 25 nonsynonymous substitutions per site per 10(9) years). cDNA encoding the mature protein (89-100 residues) shows evidence of positive selection (Dn/Ds > 1) for 4 of 10 pairwise species comparisons. cDNA and predicted secondary-structure comparisons suggest that TMAP is neither orthologous nor paralogous to abalone lysin, and thus marks a second, phylogenetically independent, protein subject to strong positive selection in free-spawning marine gastropods. In addition, an internal repeat in one species (Tegula aureotincta) produces a duplicated cleavage site which results in two alternatively processed mature proteins differing by nine amino acid residues. Such alternative processing may provide a mechanism for introducing novel amino acid sequence variation at the amino-termini of proteins. Highly divergent TMAP N-termini from two other tegulines (Tegula regina and Norrisia norrisii) may have originated by such a mechanism.

MHC class I genes in a New World primate, the cotton-top tamarin (Saguinus oedipus), have evolved by an active process of loci turnover.

Lymphocytes of a New World primate, the cotton-top tamarin (Saguinus oedipus), express classical G-related major histocompatibility complex (MHC) class I molecules with unusually limited polymorphism and variability. Three G-related loci, an F locus, an E locus, and two pseudogenes (So-N1 and So-N3) have been identified by cDNA library screening and extensive PCR analysis of both cDNA and genomic DNA from the cotton-top tamarin. Furthermore, each genus of the subfamily Callitrichinae (tamarins and marmosets) appears to express its own unique set of MHC class I genes, likely due to a rapid turnover of loci. The rapid emergence of unique MHC class I genes in the Callitrichinae genera, resulting from an active process of duplication and inactivation of loci, may account for the limited diversity of the MHC class I genes in the cotton-top tamarin. To determine the nature of the entire complement of MHC class I genes in the cotton-top tamarin, we synthesized a genomic DNA library and screened it with MHC class I-specific probes. We isolated nine new MHC class I pseudogenes from this library. These newly isolated tamarin G-related MHC class I pseudogenes are not closely related to any of their functional counterparts in the tamarin, suggesting that they do not share a recent common ancestral gene with the tamarin's currently expressed MHC class I loci. In addition, these tamarin sequences display a high rate of nonsynonymous substitutions in their putative peptide binding region. This indicates that the genes from which they have derived were likely subject to positive selection and, therefore, were once functional. Our data support the notion that an extremely high rate of loci turnover is largely responsible for the limited diversity of the MHC class I genes in the cotton-top tamarin.

Immune Avoidance Strategies in RNA Viruses: Fitness Continuums arising from Trade-offs between Immunogenicity and Antigenic Variability

Highly exposed and protruding amino acid sites on the surface of viral capsids are subject to fewer residue interactions and packing constraints than those buried within protein interiors. Consequently they often experience higher rates of non-synonymous substitution and exhibit greater genetic variability than buried interior sites. However such protrusive surface structures often induce host immune responses and are likely to constitute B cell epitopes. Genetic variation at these surface sites is therefore likely to correspond to antigenic variation. This may be of adaptive value to the virus for two quite different reasons. The first is that antigenic variation arising over the course of a viraemia may result in greater net viral replication, and increased opportunities for viral transmission. The second is that antigenic variation generated rapidly over a single infection or incrementally over several sequential infections may give rise to variants that are sufficiently immunologically distinct that they can reinfect host individuals with previous infection experience of related virus. This would lead to an extension of the susceptible host pool with consequent increase in transmission opportunities.The surface architecture of viral capsid proteins is therefore conceivably subject to two opposing selection pressures: one to minimize the surface area accessible to interactions with elements of the immune system, the other to increase the potential access to antigenic variation by adoption of exposed and unconstrained protein conformations. Therefore, there exists a possible trade-off between the fitness benefits deriving from potential ability to generate antigenic variation, and the increased immunogenicity with such potential may be associated. We propose that the existence of this trade-off would lead to a continuum of different strategies by which a virus might combat an immune response. We explore this strategy space with simple mathematical models, and show that peak loads of infectious viral particles are proportional to levels of antigenic diversity, and inversely proportional to immunogenicity, thereby creating the potential for a trade-off by which fitness might be maintained with a continuum of strategies. This may remain possible even if the antigenic variants are not transmissible between hosts, so long as immune resources are sufficiently dispersed between antigenic variants, The diversity of possible stratagies is discussed with reference to the Picornavirus family.