Institute of Molecular Evolutionary Genetics and Department of Biology, Pennsylvania State University
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Abstract |
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Key Words: MHC class I divergence time birth-and-death evolution primates
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Introduction |
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Both class I and class II MHC gene families include a large number of loci and have been shown to evolve according to the birth-and-death process (Nei and Hughes 1992; Klein et al. 1993; Nei, Gu, and Sitnikova 1997). In this process new genes are created by repeated gene duplications, and some genes may later become pseudogenes or even be deleted from the genome. As a result of the birth-and-death evolution, these multigene families consist of a mixture of divergent genes, some of which have remained in the genome for a long time, and a large number of closely related genes or pseudogenes (Ota and Nei 1994; Nei, Gu, and Sitnikova 1997). A relatively slow rate of birth-and-death evolution in class II loci makes it an attractive set of genes to study the divergence times of various loci in mammals (Klein and Figueroa 1986; Hughes and Nei 1990; Takahashi, Rooney, and Nei 2000). The longevity of these loci is relatively high; it has been estimated that most MHC class II loci originated at least 170200 MYA (Takahashi, Rooney, and Nei 2000). In contrast, it appears that class I loci experience a much faster rate of birth-and-death evolution than class II loci (Nei and Hughes 1992). As a result, there seem to be no orthologous relationships of different class I loci among different mammalian orders (Klein and Figueroa 1986; Hughes and Nei 1989). Furthermore, the divergence of class I genes occurred so recently that even humans and New World monkeys, which diverged only about 3335 MYA, do not share functional genes (Watkins et al. 1990a; Cadavid et al. 1997). Similarly, class I genes from two marsupial species, separated about 48 MYA, show no orthologous relationships (Houlden, Greville, and Sherwin 1996). Therefore, the turnover rate of these class I loci must be very high. However, no serious attempts have been made to infer the divergence times of these rapidly evolving loci, in part because there were not enough DNA sequence data. Recently a fair amount of sequence data has accumulated on the class I genes in higher primates, so it is now possible to evaluate the times of birth and death of several important MHC class I genes. Furthermore, analysis of genes from closely related species such as primates allows us to focus on relatively recent events of loci origin and divergence even within relatively fast-evolving multigene families. Here, we estimate the divergence time among primate MHC class I loci, using two phylogenetic approaches: the linearized tree method and the distance regression method.
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Materials and Methods |
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Phylogenetic Analysis
Phylogenetic analysis was conducted using the Neighbor-Joining (NJ) tree-building method (Saitou and Nei 1987) as implemented with the computer program MEGA2 (Kumar et al. 2001). Because the extent of sequence divergence was relatively small and no strong transition-transversion bias was detected, the evolutionary distances between sequences were estimated using the Jukes-Cantor (JC) distance (Jukes and Cantor 1969). Complete nucleotide sequences of all three extracellular domains (1,
2,
3) were used. All three codon positions were used. Gaps were removed from the computations using the complete-deletion option.
Two data sets were used, consisting of Platyrrhini (i.e., New World monkeys) and Catarrhini (i.e., humans, Old World monkeys, and apes) sequences. To avoid errors associated with insufficient taxon sampling (De Rijk et al. 1995; Murphy et al. 2001), human, chimpanzee, and gorilla class I sequences were used as representatives of Catarrhini species in the first data set, later referred to as the platyrrhine data set. Similarly, the second data set, later referred to as the catarrhine data set, includes sequences from two species of tamarins (S. oedipus and S. fuscicollis) as representatives of Platyrrhini clade. A total of 274 and 270 codons were used in the platyrrhine and catarrhine data sets, respectively.
The reliability of tree topologies was evaluated by the bootstrap interior branch test (Felsenstein 1985) with 500 replications, and bootstrap probability values greater than 80% were regarded as statistically significant (Sitnikova, Rzhetsky, and Nei 1995; Nei and Kumar 2000).
To examine the reliability of NJ topologies, we also constructed maximum-parsimony (MP) and maximum likelihood (ML) phylogenetic trees using the beta-version (4.0b10) of the computer program PAUP* (Swofford 2002). For each data set MP trees were generated using a heuristic search option with 10 random stepwise-addition (SA) replicates that were followed by tree bisection-reconnection (TBR) branch swapping to completion. To estimate relative branch support, bootstrap analysis (Felsenstein 1985) with 500 replicates was conducted (i.e., 500 bootstrap replications of 10SA + TBR searches). We constructed and compared 50% majority rule consensus MP trees with the NJ trees.
Maximum likelihood tree searches were conducted with the Jukes-Cantor model of nucleotide substitutions (Jukes and Cantor 1969). Reconstruction of a ML tree usually involves extensive computational efforts. However, it has been demonstrated that the most extensive search algorithm does not necessarily produces the best results (Nei, Kumar, and Takahashi 1998; Takahashi and Nei 2000) and that the combination of the bootstrap test with relatively simple search algorithms can be as efficient as more extensive searches. Here, we used a relatively simple branch swapping algorithm such as nearest-neighbor interchange (NNI) combined with the bootstrap test, rather than the extensive TBR search. The heuristic search consisted of 10 random SA followed by NNI branch swapping (i.e., 10SA + NNI). We produced 50% majority rule consensus ML trees on the basis of 100 bootstrap replications.
Divergence Time Estimation
The branch length test as implemented in the computer program LINTREE (Takezaki, Rzhetsky, and Nei 1995) was used to test the rate constancy among sequences. This test allows identifying the sequences whose evolutionary rate significantly deviates from the average rate. Following the example of Takahashi, Rooney, and Nei (2000), we used a relatively high level of significance of 0.5% to identify such deviant sequences (sees asterisks in figure 2A and B). As shown previously (Nei and Kumar 2000), even if the molecular clock assumption is violated to some extent, it is still possible to obtain reasonable time estimates. Therefore, we estimated divergence time using (1) the complete set of sequences available and (2) only those sequences that do not violate the molecular clock assumption at the 0.5% significance level. The two data sets led to similar time estimates; therefore only the results based on the complete set of sequences in each data set are presented here (see Results).
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The regression model considers dAB= 2rt, where dAB represents the average evolutionary distance between two gene clusters A and B, and t represents the divergence time associated with dAB (Hughes and Nei 1990). The evolutionary rate was estimated to be r = 1.86 x 10-9 and 1.6 x 10-9 substitutions per site per million years for the platyrrhine and catarrhine data sets, respectively (JC distance for all codon positions was used).
Supplementary Material is available online through the Nei lab's databases maintained at http://mep.bio.psu.edu/databases/MHC_I/.
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Results |
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A phylogenetic tree of Catarrhini sequences was also constructed (fig. 1B). To avoid taxon sampling errors, sequences of two species of tamarins (S. oedipus and S. fuscicollis) were used as representatives of Platyrrhini clade. As in the case of the platyrrhine data set (fig. 1A), the F locus genes of human, chimpanzee, and macaque constitute the most basal part of the phylogenetic tree. Other loci comprise separate clusters, with the classical C locus being the part of the major B locus genes cluster, confirming their common ancestry (Adams, Thomson, and Parham 1999). Furthermore, most class I genes from tamarins form a separate cluster with the human nonclassical G locus (Watkins et al. 1990a; Cadavid et al. 1997) and not with the human classical loci A, B, and C. The recently described chimpanzee-specific locus AL (Adams, Cooper, and Parham 2001) is clustered with orangutan A locus sequences (sequences of pygmy chimp.5 and pygmy chimp.6, respectively), although the bootstrap support for this group is rather low. Overall, this phylogeny indicates that while the origin of Old World monkey loci, with the exception of locus C, predated the Platyrrhini-Catarrhini divergence, the expansion of class I genes in New World monkey species occurred after the Platyrrhini-Catarrhini split.
Essentially the same tree topologies were obtained when only the first and second codon positions were used, although the bootstrap support for the majority of gene clusters decreased significantly (results not shown). Such decrease in bootstrap support can be attributed to the decrease in the number of sites involved in the resampling, arguing for the employment of as many nucleotide sites as possible (Nei and Kumar 2000).
Comparison of NJ, ML, and MP Topologies
For each data set the 50% majority rule consensus MP and ML trees were constructed (results not shown). Both the MP and ML trees showed essentially the same clustering pattern when compared with each other and with the appropriate NJ topology. The sequence clusters that received relatively high bootstrap support (80% and higher) with the NJ method were also significantly supported by the bootstrap values on the MP and ML trees. For example, the cluster confirming the common origin of B and C loci (Adams, Thomson, and Parham 1999) was supported by bootstrap values ranging from 95% to 97% on the MP trees, and 93% to 97% in ML trees. Similarly, the cluster of the F locus sequences received 99% to 100% bootstrap support in all trees examined. However, resolution of the deep divergence branches specifying the branching order of the major loci was relatively poor under the MP and ML criteria. Bootstrap support values of these branches did not exceed 50%; therefore several of these short internal branches were collapsed in 50% majority rule consensus trees. However, the lack of resolution of these branches was observed in all three methods (i.e., NJ, MP, and ML).
Because of the excessive amount of computational time, only 20 bootstrap replications of the extensive ML heuristic search (10 SA replicates followed by the TBR search) were performed. The resulting trees were then compared with the consensus trees built after 100 bootstrap replications of the less extensive search (10 SA + NNI). Overall, both heuristic procedures resulted in topologies that showed clustering patterns consistent with the appropriate NJ topologies. It appeared that use of the more extensive search, in this case TBR, does not find a better-resolved topology than the use of the simpler search algorithm, in this case NNI. Rather, both heuristic searches produced similar patterns of clustering. Furthermore, bootstrap support of the appropriate branches in NNI-based and TBR-based trees was similar, and identical to the bootstrap support of the same branches found on the NJ trees. Similar results had been observed earlier on the simulated sequence data (Nei, Kumar, and Takahashi 1998; Takahashi and Nei 2000), where use of the most extensive search algorithm cannot guarantee identification of the true tree.
Divergence Time Estimates (Linearized Trees and Distance Regression Method)
Linearized trees are presented in figure 2 (fig. 2A for platyrrhine data sets and fig. 2B for catarrhine data sets). Using the branch length test (Takezaki, Rzhetsky, and Nei 1995), a total of 7 and 12 sequences that evolve significantly slower or faster than the average at the 0.5% level were found in the platyrrhine and catarrhine data sets, respectively (these sequences are marked with asterisks in figure 2A and B). However, estimates of divergence time for major branching points of the tree constructed using only sequences that do not violate the molecular clock assumption, were found to be quite similar to those obtained using the complete set of sequences (Nei and Kumar 2000; Takahashi, Rooney, and Nei 2000) (see also our supplementary figure 3 available online at http://mep.bio.psu.edu/databases/MHC_I/). Therefore, here we present only the results obtained using the complete set of sequences in each data set. The time scales, presented in figure 2A and B, were derived under the assumption that the human and chimpanzee F locus genes (fig. 2A) and the human and orangutan E locus genes (fig. 2B) diverged about 6 and 13 MYA (Nei and Glazko 2002), respectively. Use of other CPs, including more ancient divergences, resulted in similar time scales. Numerical results of both the linearized tree method and the distance regression method are presented in table 2. Notably, divergence time estimates obtained using these two methods are close to each other for each particular data set. Furthermore, estimates of the particular divergence times, derived from different data sets, are also very close to each other.
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We also estimated the age of the putatively oldest allelic lineages for human MHC class I genes. The divergence times of the major monophyletic allelic lineages (Gu and Nei 1999) were estimated as 1419, 1015, and 1317 MYA for the putatively oldest alleles from classical class I loci A, B, and C in humans, respectively. These results are similar to those obtained by Klein, Sato, and O'HUigin (1998), suggesting that even though the B locus is presumably older than the C locus, the divergence of its oldest allelic lineage might have occurred later than the divergence of the oldest allelic lineages of loci A and C. Furthermore, analysis by Gu and Nei (1999) showed that although the B locus is the most polymorphic of the three loci (i.e., has the highest number of alleles among all three loci), these alleles can only be separated into three groups. The largest group was comprised of several clusters; however, the bootstrap support of such allelic clusters was rather low (Gu and Nei 1999). Thus the relatively high rate of interallelic recombination, observed at this locus (Watkins et al. 1992; Marcos et al. 1997), may affect the longevity of individual alleles at the B locus, reducing the putative age of individual alleles. Alleles from human nonclassical loci E and G appeared to have diverged much later, at about 0.71.3 MYA (see fig. 2).
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Discussion |
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The active role that interlocus recombination might have played in shaping the evolution of MHC genes has been discussed extensively (Pease et al. 1991; Hogstrand and Bohme 1994; Yun, Melvold, and Pease 1997). In this case, the genes from several loci within species are expected to be more similar to each other than to the genes from the other species. However, our results show that the genes from closely related species such as human and chimpanzee do not form species-specific clusters. Instead, these genes cluster in a loci-specific manner (see fig. 1). Similarly, genes from two tamarin species are intermingled on a tree, indicating that these genes have maintained their genetic identity since the speciation event. Therefore, the potential role that genetic exchanges such as gene conversion and unequal crossing-over may play in the long-term evolution of MHC genes in primates appears to be rather small. Similarly, analyzing human and mouse MHC genes, Gu and Nei (1999) showed that the overall frequency of interlocus recombination is small and that most of the genetic variation observed between MHC loci should be attributed to selection and mutation.
Our divergence time estimates suggest that some of MHC class I loci, in particular the F locus, were maintained in the primate genome for a long time, apparently at least 4666 My. Other loci, such as A, B, G, and E, were estimated to have originated slightly later, at around 3946 MYA, in the time period that predates the Catarrhini-Platyrrhini clade split (Adams, Cooper, and Parham 2001). There are also examples of more recent gene duplications leading to the appearance of new MHC class I loci. In particular, chimpanzee AL and duplicate B loci in macaque were estimated to have originated around 1825 and 2331 MYA, respectively (table 2), which makes them relatively young loci. Overall, our divergence time estimates are falling in agreement with the molecular estimates of the primate speciation dates (Goodman et al. 1998; Nei and Glazko 2002).
At present the question of whether some Platyrrhini species possess genes homologous to the classical loci of Catarrhini remains unclear. In particular, genes Ateles B*01 and Pithecia B*01 from spider monkey and saki, respectively, clustered significantly with the other B and C loci sequences (Watkins 1995; Cadavid et al. 1997) suggesting their orthologous relationship. In our analysis, these alleles (designated spider monkey.2 and saki.4, respectively) do not cluster with the B and C loci. Instead, they form a separate cluster within other platyrrhine sequences, although the bootstrap support value for the internal branch was low (below 50%). Furthermore, this cluster takes a basal position among other G-locusrelated Platyrrhini sequences, but again the bootstrap support for that pattern is rather low (see fig. 1A). Similarly, Adams and Parham (2001) observed that these two genes were not clustered phylogenetically any closer with the classical B locus than they were with the nonclassical G locus genes.
Currently, the exact order of appearance of class I loci in primates remains unclear. On the basis of the genomic structure of the MHC region in humans, it has been suggested that the duplication of the F locus led to the appearance of the G locus (Shiina et al. 1999) before other loci, such as A, B, and E, arose. The putative orthologous relationships that we observe among the locus G of great apes and humans and the appropriate loci of platyrrhine species (Watkins et al. 1990b; Cadavid et al. 1997; Adams and Parham 2001) may be used to support this idea. Our results suggest that loci A, B, and E originated rather quickly after the gene duplication that gave rise to locus G. Furthermore, our estimates place the time of origin of these major class I loci somewhat before the time of the Catarrhini-Platyrrhini divergence. These findings, together with the possibility that particular MHC genes have persisted in the genome for a long time, suggest that some platyrrhine species may still retain genes, orthologous to the classical A and B loci of catarrhine species. Some studies suggest that such genes may indeed exist (Watkins et al. 1990a; Watkins 1995; Cadavid et al. 1997), but the final answer to this question will come from genomic studies involving the analysis of complete genomic sequences of many primate species as well as other mammals. Current advances in complete genome sequencing and mapping should enable us to further study the evolutionary dynamics of the MHC region in a variety of organisms and to better understand the evolutionary factors affecting MHC evolution.
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Acknowledgements |
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Footnotes |
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Literature Cited |
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Adams, E. J., S. Cooper, and P. Parham. 2001. A novel, nonclassical MHC class I molecule specific to the common chimpanzee. J. Immunol. 167:3858-3869.
Adams, E. J., and P. Parham. 2001. Species-specific evolution of MHC class I genes in the higher primates. Immunol. Rev. 183:41-64.[CrossRef][ISI][Medline]
Adams, E. J., G. Thomson, and P. Parham. 1999. Evidence for an HLA-C-like locus in the orangutan Pongo pygmaeus. Immunogenetics 49:865-871.[CrossRef][ISI][Medline]
Bjorkman, P. J., M. A. Saper, B. Samraoui, W. S. Bennett, J. L. Strominger, and D. C. Wiley. 1987. The foreign antigen binding site and T cell recognition regions of class I histocompatibility antigens. Nature 329:512-518.[CrossRef][ISI][Medline]
Boyson, J. E., C. Shufflebotham, L. F. Cadavid, J. A. Urvater, L. A. Knapp, A. L. Hughes, and D. I. Watkins. 1996. The MHC class I genes of the rhesus monkey. Different evolutionary histories of MHC class I and II genes in primates. J. Immunol. 156:4656-4665.
Cadavid, L. F., A. L. Hughes, and D. I. Watkins. 1996. MHC class I-processed pseudogenes in New World primates provide evidence for rapid turnover of MHC class I genes. J. Immunol. 157:2403-2409.[Abstract]
Cadavid, L. F., C. Shufflebotham, F. J. Ruiz, M. Yeager, A. L. Hughes, and D. I. Watkins. 1997. Evolutionary instability of the major histocompatibility complex class I loci in New World primates. Proc. Natl. Acad. Sci. USA 94:14536-14541.
De Rijk, P., Y. Van de Peer, I. Van den Broeck, and R. De Wachter. 1995. Evolution according to large ribosomal subunit RNA. J. Mol. Evol. 41:366-375.[ISI][Medline]
Dixon, B., and R. J. Stet. 2001. The relationship between major histocompatibility receptors and innate immunity in teleost fish. Dev. Comp. Immunol. 25:683-699.[CrossRef][ISI][Medline]
Felsenstein, J. 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39:783-791.[ISI]
Goodman, M., C. A. Porter, J. Czelusniak, S. L. Page, H. Schneider, J. Shoshani, G. Gunnell, and C. P. Groves. 1998. Toward a phylogenetic classification of primates based on DNA evidence complemented by fossil evidence. Mol. Phylogenet. Evol. 9:585-598.[CrossRef][ISI][Medline]
Gu, X., and M. Nei. 1999. Locus specificity of polymorphic alleles and evolution by a birth-and-death process in mammalian MHC genes. Mol. Biol. Evol. 16:147-156.[Abstract]
Harland W. B., R. L. Armstrong, A. V. Cox, L. E. Craig, A. G. Smitch, and D. G. Smith. 1990. A geologic time scale. Cambridge University Press, Cambridge.
Hogstrand, K., and J. Bohme. 1994. A determination of the frequency of gene conversion in unmanipulated mouse sperm. Proc. Natl. Acad. Sci. USA 91:9921-9925.
Houlden, B. A., W. D. Greville, and W. B. Sherwin. 1996. Evolution of MHC class I loci in marsupials: characterization of sequences from koala (Phascolarctos cinereus). Mol. Biol. Evol. 13:1119-11127.[Abstract]
Hughes, A. L. 1995. Origin and evolution of HLA class I pseudogenes. Mol. Biol. Evol. 12:247-258.[Abstract]
Hughes, A. L., and M. Nei. 1989. Evolution of the major histocompatibility complex: independent origin of nonclassical class I genes in different groups of mammals. Mol. Biol. Evol. 6:559-579.[Abstract]
1990. Evolutionary relationships of class II major-histocompatibility-complex genes in mammals. Mol. Biol. Evol. 7:491-514.[Abstract]
Jukes, T. H., and C. R. Cantor. 1969. Evolution of protein molecules. Pp. 21132 in H. N. Munro, ed. Mammalian protein metabolism. Academic Press, New York.
Klein, J., and F. Figueroa. 1986. Evolution of the major histocompatibility complex. Crit. Rev. Immunol. 6:295-386.[Medline]
Klein, J., and V. Horejsi. 1997. Immunology. Blackwell Science, London.
Klein, J., H. Ono, D. Klein, and C. O'hUigin. 1993. The accordion model of MHC evolution. Prog. Immunol. 8:137-143.
Klein, J., A. Sato, and C. O'hUigin. 1998. Molecular trans-species polymorphism. Annu. Rev. Ecol. Syst. 29:1-21.[CrossRef][ISI]
Knapp, L. A., L. F. Cadavid, and D. I. Watkins. 1998. The MHC-E locus is the most well conserved of all known primate class I histocompatibility genes. J. Immunol. 160:189-196.
Kumar, S., K. Tamura, I. B. Jakobsen, and M. Nei. 2001. MEGA2: molecular evolutionary genetics analysis software. Bioinformatics 17:1244-1245.
Lawlor, D. A., E. Warren, F. E. Ward, and P. Parham. 1990. Comparison of class I MHC alleles in humans and apes. Immunol. Rev. 113:147-185.[ISI][Medline]
Marcos, C. Y., M. A. Fernandez-Vina, A. M. Lazaro, C. J. Nulf, E. H. Raimondi, and P. Stastny. 1997. Novel HLA-B35 subtypes: putative gene conversion events with donor sequences from alleles common in native Americans (HLA-B*4002 or B*4801). Hum. Immunol. 53:148-155.[CrossRef][ISI][Medline]
Murphy, W. J., E. Eizirik, W. E. Johnson, Y. P. Zhang, O. A. Ryder, and S. J. O'Brien. 2001. Molecular phylogenetics and the origins of placental mammals. Nature 409:614-618.[CrossRef][ISI][Medline]
Nei, M., and G. V. Glazko. 2002. The Wilhelmine E. Key 2001 Invitational Lecture. Estimation of divergence times for a few mammalian and several primate species. J. Hered. 93:157-164.
Nei, M., X. Gu, and T. Sitnikova. 1997. Evolution by the birth-and-death process in multigene families of the vertebrate immune system. Proc. Natl. Acad. Sci. USA 94:7799-7806.
Nei, M., and A. L. Hughes. 1992. Balanced polymorphism and evolution by the birth-and-death process in the MHC loci. Pp. 2738 in K. Tsuji, M. Aizawa, and T. Sasazuki, eds. 11th Histocompatibility Workshop and Conference. Oxford University Press, Oxford.
Nei, M., and S. Kumar. 2000. Molecular evolution and phylogenetics. Oxford University Press, Oxford.
Nei, M., S. Kumar, and K. Takahashi. 1998. The optimization principle in phylogenetic analysis tends to give incorrect topologies when the number of nucleotides or amino acids used is small. Proc. Natl. Acad. Sci. USA 95:12390-12397.
Ota, T., and M. Nei. 1994. Divergent evolution and evolution by the birth-and-death process in the immunoglobulin VH gene family. Mol. Biol. Evol. 11:469-482.[Abstract]
Otting, N., and R. E. Bontrop. 1993. Characterization of the rhesus macaque (Macaca mulatta) equivalent of HLA-F. Immunogenetics 38:141-145.[ISI][Medline]
Pease, L. R., R. M. Horton, J. K. Pullen, and Z. L. Cai. 1991. Structure and diversity of class I antigen presenting molecules in the mouse. Crit. Rev. Immunol. 11:1-32.[ISI][Medline]
Saitou, N., and M. Nei. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol. 4:406-425.[Abstract]
Schneider, H. 2000. The current status of the New World monkey phylogeny. An. Acad. Bras. Cienc. 72:165-172.[ISI][Medline]
Shiina, T., G. Tamiya, and A. Oka, et al. (23 coauthors). 1999. Molecular dynamics of MHC genesis unraveled by sequence analysis of the 1,796,938-bp HLA class I region. Proc. Natl. Acad. Sci. USA 96:13282-13287.
Sitnikova, T., A. Rzhetsky, and M. Nei. 1995. Interior-branch and bootstrap tests of phylogenetic trees. Mol. Biol. Evol. 12:319-333.[Abstract]
Stauffer, R. L., A. Walker, O. A. Ryder, M. Lyons-Weiler, and S. B. Hedges. 2001. Human and ape molecular clocks and constraints on paleontological hypotheses. J. Hered. 92:469-474.
Swofford, D. L. 2002. PAUP*: phylogenetic analysis using parsimony (*and other methods). Version 4. Sinauer Associates, Sunderland, Mass.
Takahashi, K., and M. Nei. 2000. Efficiencies of fast algorithms of phylogenetic inference under the criteria of maximum parsimony, minimum evolution, and maximum likelihood when a large number of sequences are used. Mol. Biol. Evol. 17:1251-1258.
Takahashi, K., A. P. Rooney, and M. Nei. 2000. Origins and divergence times of mammalian class II MHC gene clusters. J. Hered. 19:198-204.[CrossRef]
Takezaki, N., A. Rzhetsky, and M. Nei. 1995. Phylogenetic test of the molecular clock and linearized trees. Mol. Biol. Evol. 12:823-833.[Abstract]
Thompson, J. D., T. J. Gibson, F. Plewniak, F. Jeanmougin, and D. G. Higgins. 1997. The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res. 25:4876-4882.
Watkins, D. I. 1995. The evolution of major histocompatibility class I genes in primates. Crit. Rev. Immunol. 15:1-29.[ISI][Medline]
Watkins, D. I., Z. W. Chen, A. L. Hughes, M. G. Evans, T. F. Tedder, and N. L. Letvin. 1990a. Evolution of the MHC class I genes of a New World primate from ancestral homologues of human non-classical genes. Nature 346:60-63.[CrossRef][ISI][Medline]
Watkins, D. I., T. L. Garber, Z. W. Chen, G. Toukatly, A. L. Hughes, and N. L. Letvin. 1991. Unusually limited nucleotide sequence variation of the expressed major histocompatibility complex class I genes of a New World primate species (Saguinus oedipus). Immunogenetics 33:79-89.[ISI][Medline]
Watkins, D. I., N. L. Letvin, A. L. Hughes, and T. F. Tedder. 1990b. Molecular cloning of cDNA that encode MHC class I molecules from a New World primate (Saguinus oedipus). Natural selection acts at positions that may affect peptide presentation to T cells. J. Immunol. 144:1136-1143.
Watkins, D. I., S. N. McAdam, and X. Liu, et al. (13 coauthors). 1992. New recombinant HLA-B alleles in a tribe of South American Amerindians indicate rapid evolution of MHC class I loci. Nature 357:329-333.[CrossRef][ISI][Medline]
Yun, T. J., R. W. Melvold, and L. R. Pease. 1997. A complex major histocompatibility complex D locus variant generated by an unusual recombination mechanism in mice. Proc. Natl. Acad. Sci. USA 94:1384-1389.