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MBE Advance Access originally published online on September 14, 2005
Molecular Biology and Evolution 2006 23(1):137-143; doi:10.1093/molbev/msj016
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Research Article

Evolution by Recombination and Transspecies Polymorphism in the MHC Class I Gene of Xenopus laevis

David H. Bos1 and Bruce Waldman

School of Biological Sciences, University of Canterbury, Christchurch, New Zealand

E-mail: dbos{at}purdue.edu.


   Abstract
TOP
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Acknowledgements
 References
 
The patterns of major histocompatibility complex (MHC) evolution involve duplications, deletions, and independent divergence of loci during episodes punctuated by natural selection. Major differences in MHC evolution among taxa have previously been attributed to variation in linkage patterns of class I and class II MHC genes. Here we characterize patterns of evolution in the MHC class Ia gene of Xenopus laevis in terms of polymorphism, recombination, and extent of transspecies polymorphism. We also compare these patterns to see if a correlation exists with linkage or separation of the MHC class I and class II regions as seen in amphibians and teleost fishes. In X. laevis, we find high levels of polymorphism. Also, genetic exchange is relatively frequent and occurs in intron II, reshuffling allelic forms of exons 2 and 3. Evolutionary relationships among class I alleles show an intermingling of alleles from divergent Xenopus species rather than a species-specific clustering. Results indicate that the patterns of evolution are similar to those found in salmonid fishes and are different from the mode of evolution seen in primates. Similar patterns of class Ia evolution in salmonid fishes and X. laevis suggest that nonlinkage of class I and class II regions alone is insufficient to explain some patterns of MHC evolution in salmonids.

Key Words: transspecies evolution • recombination • exon shuffling • Akaike information criterion (AIC) • Xenopus • MHC


   Introduction
TOP
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Acknowledgements
 References
 
Major histocompatibility complex (MHC) class I and class II genes are found in all gnathostomes and encode structurally similar proteins that present antigenic peptides to T lymphocytes. Class II proteins are expressed mainly on specialized antigen-presenting cells and primarily function to bind peptides derived from extracellular pathogens. Class I proteins are expressed in almost all cells and are involved in monitoring the internal environment of the cell for foreign, mutated, or misfolded proteins. Class I genes comprise classical (class Ia) and nonclassical (class Ib) loci which differ in polymorphism, structure, function, and expression pattern. Aside from their important role in the immune system, the MHC genes are of particular interest because of their patterns of genetic diversity.

Class I and class II alleles have several exceptional features. They exhibit very high levels of allelic diversity and amino acid polymorphism (Parham and Ohta 1996Go). Some MHC allelic lineages also exhibit unusual longevity which predates the formation of species (Figueroa, Gunther, and Klein 1988Go; Lawlor et al. 1988Go). As a result, "transspecies polymorphisms" exist, whereby some MHC alleles from separate species cluster together in phylogenetic analysis to the exclusion of alleles from within the same species. Variations in linkage patterns, order of gene loci, and the number of gene family members resulting from tandem duplications have also been observed (Kelley, Walter, and Trowsdale 2005Go). Many of the features of the MHC have been attributed to the forces of balancing selection acting at the molecular level (Hughes and Yeager 1998Go).

In the typical pattern of MHC evolution, class I and class II families of genes evolve differently from each other, each with its own rate of duplication, divergence, longevity, and pattern of recombination. Specifically, class II transspecies polymorphisms extend further back in evolutionary history than in class I lineages (Bontrop et al. 1999Go; Vogel et al. 1999Go). Class II loci also have higher levels of polymorphism than class I loci. Additionally, allelic recombination has been characterized by the intralocus exchange of small minicassettes of nucleotides that can occur throughout the length of the gene and has no single prominent breakpoint (A. L. Hughes, M. K. Hughes, and Watkins 1993Go; Jakobsen, Wilson, and Easteal 1998Go). Until more recently, it was not known whether these well-established patterns of MHC evolution were also found in species other than the mammalian model organisms.

Despite many elements of conserved structure and function, MHC evolution in nonmammalian taxa differs from well-defined norms. For instance, class II gene evolution in birds differs from that in mammals; the loci have a relatively recent origin in birds (Edwards, Wakeland, and Potts 1995Go; Hess and Edwards 2002Go). In bony fishes, the class I and class II genes, linked in a common region in all other vertebrates, are found on separate chromosomes (Bingulac-Popovic et al. 1997Go). However, this appears to be a derived trait in bony fishes because both class I and class II genes of sharks are closely linked (Ohta et al. 2000Go). Class Ia lineages in salmonid fishes share transspecies polymorphism among divergent taxa, whereas class II alleles cluster in a species-specific manner, just opposite the pattern in mammalian model organisms (Shum et al. 2001Go). Salmonid fishes also have much higher levels of class Ia than class II polymorphism. In general, recombination plays a more prominent role in teleost class Ia evolution than in mammalian benchmarks (Shum et al. 2001Go; Consuegra et al. 2005Go). Intragenic recombination in salmonids typically involves entire exons, and a prominent breakpoint for genetic exchange is easily identifiable. Authors have speculated that these patterns of evolution might be due to the nonlinkage of class I and class II loci as seen in bony fishes (Shum et al. 2001Go).

Xenopus laevis, the African clawed frog, is the first ectothermic vertebrate from which class I proteins were isolated (Flajnik et al. 1984Go). In this species, there is a single MHC class Ia locus with diploid inheritance patterns (Shum et al. 1993Go). This locus is highly polymorphic compared to mammals but is similar to the variation found in salmonid fishes. Another unusual aspect of Xenopus class Ia is the existence of two ancient allelic lineages (Flajnik et al. 1999Go). These lineages are very distinct as alleles belonging to different lineages are as divergent as MHC alleles from mouse and human. Linkage of class Ia and class II genes in X. laevis indicates a single MHC genomic region like many vertebrates, but unlike bony fishes (Nonaka et al. 1997Go).

Our motivation is to ascertain patterns of MHC class I evolution, by characterizing recombination, polymorphism, and extent of transspecies evolution in the class Ia gene of wild-caught Xenopus frogs. Previously established differences in the linkage of class I and class II regions between Xenopus and salmonid fishes also allow us to compare results and interpret these in light of hypotheses regarding the influence of linkage on MHC evolution. Differing patterns of class Ia evolution among these taxa would support the hypothesis that the mode of MHC evolution seen in salmonid fishes might be due to the nonlinkage of class I and class II genes. This mode of evolution was reported by Shum et al. (2001)Go and is distinguished by ancient class I lineages, high levels of polymorphism, and frequent recombination between the peptide-binding region (PBR)–coding exons. However, class Ia evolution in X. laevis that is similar to that of fishes supports the notion that nonlinkage of class Ia and class II genes alone may be insufficient to explain the mode of class Ia evolution reported for salmonids.


   Materials and Methods
TOP
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Acknowledgements
 References
 
Data Collection
We extracted total RNA from X. laevis blood samples using the TRIzol protocol following the manufacturer's recommendations (Invitrogen). Total RNA was used to make cDNA in a Superscript One-Step reverse transcriptase–polymerase chain reaction (PCR) kit (Invitrogen, Carlsbad, CA), and first-strand synthesis was performed at 55°C for 25 min. Immediately after the first-strand synthesis, PCR was employed on cDNA with primers designed to amplify exons 2–4 of the MHC class Ia gene (forward primer: 5'-GTCACTCCCTGCGYTAYTAT-3' and reverse primer: 5'-TTTCTCCTTCAGGCTGCTGT-3'). Primers were designed using Primer3 (Rozen and Skaletsky 2000Go) from known X. laevis sequences (Flajnik et al. 1999Go). The thermal profile used to amplify MHC fragments was optimized to minimize the occurrence of in vitro recombination (Judo, Wedel, and Wilson 1998Go). We cloned PCR products into the pCR 4 TOPO TA plasmid following the manufacturer's recommendations (Invitrogen), and recombinant DNA was transformed into TOP-10 Escherichia coli cells. Escherichia coli cells were plated onto Luria-Bertani (LB) agar and grown overnight at 37°C after which 6–10 individual colonies were picked and grown in LB broth at 37°C for 16 h.

A total of 5 ml of LB broth per cell matrix was removed, and plasmid DNA was extracted using alkaline lysis minipreps (Sambrook, Fritsch, and Maniatis 1989Go). We sequenced the MHC insert in both directions using BigDye v3.1 chemistry and an ABI 3730 automated sequencer. ABI trace files were edited using Bioedit (Hall 1999Go), and sequences were aligned using ClustalW (Thompson, Higgins, and Gibson 1994Go). Eleven new sequences were isolated from 11 X. laevis chromosomes; these sequences were independently verified from two to six separate colonies (GenBank accession numbers: DQ149596–DQ149606). Additional sequences were obtained but were not recovered multiple times and were excluded from the following analyses. New sequences were added to other known class Ia sequences of exons 2, 3, and 4 from frogs (GenBank accession numbers—Xenopus tropicalis: AY204558, AY204559 [X. tropicalis is also termed Silurna tropicalis but is listed herein as a congeneric Xenopus species due to strong monophyly of this species with other Xenopus {Evans et al. 2004Go}]; Xenopus ruwenzoriensis: AF497525–AF497528; and X. laevis, Rana pipiens, and a laboratory-bred interspecies hybrid of X. laevis-Xenopus gilli: AF185579–AF185588).

Statistical Analysis
We used various statistical methods to investigate evolutionary relationships and intragenic recombination. The program Maxchi was used to detect the occurrence of recombination events within X. laevis samples because it performs well in simulations and had a low false error rate (Posada and Crandall 2001cGo). The program RDP2 (Martin, Williamson, and Posada 2005Go) characterized intragenic recombination by identifying breakpoints and alleles created by recombination. The P value of significant differences used to infer recombination was set at 0.000005 with a window size of 20 nt to minimize the false-positive error rate (Martin, Williamson, and Posada 2005Go). To estimate population parameters (i.e., mutation [{theta} = 4Nµ] and recombination [{rho} = 4Nr]), we used the maximum likelihood (ML) method implemented in LDhat because this method uses a finite-sites model appropriate for highly divergent sequences (McVean, Awadalla, and Fearnhead 2002Go).

Evolutionary relationships were reconstructed among known Xenopus MHC class Ia sequences. Prior to phylogenetic analysis, we tested for the loss of information in these data due to saturation using the index of substitution saturation (ISS) (Xia et al. 2003Go). We compared these values with the symmetric critical index of substitution saturation (ISS.C) scores. For this procedure, we estimated the proportion of invariable sites and tested each data partition (see below) separately; each test included all sequences (n = 27). We used model-based algorithms to investigate evolutionary relationships among sequences (Bos and Posada 2005Go). The best approximating model of nucleotide evolution for these data was determined using Akaike's information criterion (AIC) (Akaike 1974Go). ML scores of candidate models were calculated using PAUP* 4.0 (Swofford 1998Go) and AIC scores computed in Modeltest (Posada and Crandall 1998Go). Employing the best approximating model, genetic distance and phylogenetic relationships were estimated using ML optimization.

A traditional bifurcating phylogenetic tree may not accurately represent evolutionary relationships among a population sample because of genetic exchange (Posada and Crandall 2001aGo). Therefore, we reconstructed separate trees by partitioning these data into congruent segments with shared evolutionary history on either side of a putative recombination breakpoint. ML (Felsenstein 1981Go) and Neighbor-Joining (NJ; Saitou and Nei 1987Go) reconstructions were performed to test hypotheses regarding lineage assortment among species. ML trees were compared to a range of a priori topologies corresponding to different levels of transspecies lineage sharing among taxa. Tree comparisons were performed using Shimodaira-Hasegawa tests (Shimodaira and Hasegawa 1999Go; Shimodaira 2002Go) implemented in the program package Consel (Shimodaira and Hasegawa 2001Go). One thousand bootstrap pseudoreplications were used to estimate support for nodes in the NJ tree, and >50% bootstrap support in resulting topologies is shown.


   Results
TOP
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Acknowledgements
 References
 
Data
Overall, the data (including Rana outgroup samples) consist of 27 sequences and have 397 polymorphic sites out of a total of 781 nt; on average, alleles differ by 99.30 nt. The ISS.C represents the ISS value beyond which data fail to recover a true phylogenetic tree. The {alpha}1 partition is 255 bp, and the {alpha}2/{alpha}3 domain partition is 526 bp; the respective ISS values are 0.261 and 0.228. The ISS.C are 0.682 and 0.715, respectively, and these are significantly larger than the respective ISS scores (P < 0.001).

Population Parameters
Intragenic recombination plays a prominent role in X. laevis MHC evolution and is responsible for the creation of a number of new alleles. Among X. laevis sequences (including the X. laevis-X. gilli hybrids), the number of alleles created through recombination is at least 6 out of 19, over 30% of alleles in this data set (table 1). The parameters of RDP2 were set conservatively to avoid false-positive identification of recombination events, so this number represents a minimum number of recombinant alleles. Estimation of the population parameters shows relatively high mutation and recombination rates (table 2). Estimated values indicate that past mutation and recombination events both operate on the same scale and play a major role in shaping variation at this locus.


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  		Table 1 Xenopus laevis Recombinant Sequences at the UAA Locus

 

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  		Table 2 Population Parameters of Xenopus laevis Nucleotide Sequences (n = 19 samples chromosomes)

 
The recombination breakpoint also is shared among alleles, indicating that recombination is not free. The breakpoint of the recombinant alleles indicates the likely crossover location of the genetic exchange, which commonly occurs in intron II in these data. The size of the DNA fragment involved in the exchanges typically encompasses about the first 255 nt or the entire {alpha}1 domain–coding exon (tables 1 and 2). This type of recombination leads to intralocus allelic "exon shuffling" that creates new arrangements of existing variation in the PBR. This pattern is very different from that seen in humans, where genetic exchange involves much smaller fragments.

Some other trends in the pattern of recombination in X. laevis are noteworthy. For instance, some alleles are involved in genetic exchange more often than others. The occurrence of genetic exchange often involves Xela-UAA*02 in these data. This allele is involved in creating at least three new alleles; Xela-UAA*06 could be a parent sequence for two additional recombinant alleles. A bias in alleles involved in recombination has also been detected in salmonid fishes (Shum et al. 2001Go). Inspection of recombinants reveals that two recombinant alleles (alleles UAA*06 and UAA*11) have identical {alpha}1 domain sequences, but the {alpha}2 and {alpha}3 domains of these two alleles are different. Finally, the formation of recombinant alleles is not restricted to closely related alleles as two highly divergent alleles can recombine (e.g., UAAf and UAA*02). The apparent ongoing genetic exchange results in a high level of recombination that is likely to affect the evolutionary relationships among alleles and different domains of alleles.

Evolutionary Relationships
The evolutionary relationships among MHC class Ia alleles in Xenopus species were determined using ML estimation of genetic distances and NJ topology reconstruction. Tree reconstruction was done separately on two segments of the sequence, partitioning the nucleotide sequence fragment coding the {alpha}1 domain as one segment and the {alpha}2 and {alpha}3 domains together as the other segment. This partition was chosen to maximize the detection of different evolutionary histories due to genetic exchange and provides a means for confirming the presence of recombination in this data set. In the phylogenetic trees of the {alpha}1 and {alpha}2/{alpha}3 domains, recombinant alleles identified with the program RDP2 were found to be in different clades. These alleles moved across nodes with >50% bootstrap support to associate with different sets of alleles in each tree reconstruction. The translocation of alleles to different parts of the tree topology is consistent with patterns of recombination detected with RDP2.

The best approximating model selected for {alpha}1 domains differs from that chosen for the {alpha}2/{alpha}3 sequences (table 3). The best-fitting model for the {alpha}1 domain sequence evolution is (the transitional model with a gamma distribution for rates among sites TIM + {Gamma}) (Posada and Crandall 2001bGo); for the {alpha}2/{alpha}3 sequence fragment, the K81uf + {Gamma} (Kimura 1981Go) model was the best fit. Differences between data partitions are also found in tree topology, although the conservative Shimodiara-Hasagawa test indicates that the trees are not significantly different (P > 0.05).


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  		Table 3 Best Approximating Model Parameters for Data Partitions

 
The topology showing relationships among {alpha}1 domain sequences shows mixing of alleles from different species (fig. 1). Pairs of alleles from a species form well-supported groups in some cases, but both X. tropicalis and X. ruwenzoriensis {alpha}1 domains are intermingled together with X. laevis sequences. One group of X. laevis alleles is closely related and forms a tight cluster that has 100% bootstrap support; this group comprises five recombinant sequences. Most well-supported clades are near the tips of the tree and are comprised of only a few sequences; branches in the more basal parts of the tree are typically shorter than many terminal branches.



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  		FIG. 1.— Evolutionary relationships of Xenopus class Ia sequences using {alpha}1 domain sequences. Numbers indicate bootstrap support for nodes. All branches shown to the scale at the bottom left of the figure, except the branch leading to the outgroup, which was shortened for graphical clarity of the remaining branches of the tree. Rapi, Rana pipiens; Xela, Xenopus laevis; Xetr, Xenopus tropicalis; Xeru, Xenopus ruwenzoriensis; and lg, X. laevis-Xenopus gilli laboratory hybrid.

 
The evolutionary relationships reconstructed for {alpha}2/{alpha}3 domain sequences were different in some ways from the {alpha}1 domain sequence topology (fig. 2). For instance, some alleles segregated by species rather than being intermingled. In this tree, X. tropicalis alleles are basal in the topology and paraphyletic with respect to a clade of X. laevis, X. gilli, and X. ruwenzoriensis alleles. This topology establishes an incomplete separation of X. tropicalis alleles and monophyly of other sequences of the ingroup. All X. ruwenzoriensis alleles form a monophyletic cluster nested within a larger clade of X. laevis and X. gilli. There is a closely related group that forms a tight cluster similar to that seen in the {alpha}1 domain tree, but the cluster is comprised of different sequences and contains only one recombinant allele. This tree also has a mixture of both long and short terminal branches, but compared to the {alpha}1 domain tree, branches on the {alpha}2/{alpha}3 tree are much shorter.



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  		FIG. 2.— Evolutionary relationships of Xenopus class Ia sequences with Rana outgroup using {alpha}2 and {alpha}3 domain sequences. Numbers indicate bootstrap support for nodes. All branches scaled as in figure 1.

 
Several a priori hypotheses were compared to the ML and NJ topologies for both the {alpha}1 and {alpha}2/{alpha}3 sequence partitions. A priori hypotheses are not exhaustive but are designed to gauge the level of transspecies allelic sharing among progressively more distantly related species. For instance, transspecies polymorphism could be confined to two relatively closely related species, such as X. laevis and X. ruwenzoriensis, or it may extend to more distantly related taxa, such as X. laevis and X. tropicalis. Hypotheses are designed to gauge similarity of the observed level of transspecies polymorphism with constraints that represent various combinations of species that share alleles. Representative hypotheses include (1) no transspecies polymorphisms or reciprocal monophyly of species ((X. laevis),(X. ruwenzoriensis),(X. tropicalis),(R. pipiens)), (2) unconstrained X. laevis (X. laevis, (X. ruwenzoriensis),(X. tropicalis),(R. pipiens)), (3) transspecies polymorphism among two closely related species or unconstrained X. laevis and X. ruwenzoriensis (X. laevis, X. ruwenzoriensis, (X. tropicalis),(R. pipiens)), and (4) monophyletic X. laevis ((X. laevis), X. ruwenzoriensis, X. tropicalis, (R. pipiens)). Note that the ML tree serves to represent another hypothesis, namely, that transspecies polymorphism can occur throughout the genus Xenopus. Results of the Shimodaira-Hasegawa tests for both data partitions show that there is no significant difference between the ML and NJ trees (table 4). However, hypotheses 1 and 4 are significantly different from unconstrained optimal trees for both data partitions. For the {alpha}1 domain, hypothesis 2 is also significantly different from the optimal tree. For the {alpha}2/{alpha}3 data partition, tree constraints for hypotheses 2 and 3 resulted in the same phylogenetic reconstruction. Only hypothesis 3, where the only constraint is placing R. pipiens as an outgroup to Xenopus, is not significantly different from the unconstrained ML and NJ topologies for all data partitions.


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  		Table 4 Statistical Comparison of Phylogenetic Hypotheses for Xenopus Class Ia Alleles

 

   Discussion
TOP
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Acknowledgements
 References
 
Recombination
Intragenic recombination plays a prominent role in creating and maintaining diversity in the class Ia gene of X. laevis. Here, a clear pattern of reticulate evolution is characterized by high rates and phylogenetic inconsistencies between the {alpha}1 domain and sites in the remaining 3' exons of the gene. The nature of X. laevis recombination events is in contrast to the reticulate evolution seen among alleles of humans for three class I loci. Also, the pattern seen here would not be expected from in vitro chimera formation, where random breakpoints would be expected; therefore, we interpret this result to be due to in vivo allelic recombination. Although we have identified one recombination breakpoint in these sequences, the location of the other endpoint remains uncertain. However, given strong linkage disequilibrium with functionally related low molecular mass polypeptide (LMP) and transporter associated with antigen processing (TAP) loci (Namikawa et al. 1995Go; Ohta et al. 2003Go), genetic exchange events are expected to be much smaller than the distance separating these loci (Andolfatto and Nordborg 1998Go).

The high rate of genetic exchange in X. laevis could be influenced by large intron size, as seen in salmonid fishes (Shum et al. 2002Go). We attempted to PCR amplify the intron intervening exons 2 and 3 of the class Ia gene but were unable to recover any specific amplification product. However, bioinformatic exploration of a draft of the X. tropicalis genome indicates that a very large intron separates exons 2 and 3 of a class I gene. The largest intron in a class I gene was reported by Shiina et al. (2005)Go; however, the draft genome of X. tropicalis indicates that the corresponding intron is much larger (D. H. Bos, unpublished data). If intron size in a closely related species is an indication of class I intron size in X. laevis, then it is probable that recombination rates in the class I gene are influenced by these genomic features. The high recombination rate and the location of recombination events support the notion that these events may relieve possible Hill-Robertson interference (Hill and Robertson 1966Go; Otto and Barton 1997Go) on the evolution of PBR domains with differing selective pressures (Kaufman et al. 1992Go).

Phylogenetics of Xenopus MHC
Class I alleles fall into lineages that are long lasting and predate certain speciation events within the genus Xenopus. These conclusions are upheld when evolutionary relationships are reconstructed with either {alpha}1 domain sequences or {alpha}2/{alpha}3 domain sequences. However, we note that although the sequences may be useful in terms of the test of substitution saturation, our conclusions are based on partitioned data, resulting in short sequences which have less power to resolve and support relationships. In reconstructing evolutionary analysis on MHC genes, other authors have used the "total evidence" approach (Kluge 1989Go) to establish the species distribution of allelic lineages (e.g., Shum et al. 2001Go). Such an approach using ML methods on these data also confirm the above conclusions (data not shown).

Based on our reconstruction of evolutionary relationships, transspecies sharing of allelic lineages among certain divergent Xenopus species takes place. Comparison of this hypothesis with scenarios that constrain allelic separation among various species groups confirms this result. Class Ia transspecies evolution in Xenopus extends to species thought to be much more evolutionarily divergent than the species among which allele-lineage sharing is commonly found in primate class I genes. While the class Ia sequences from all Xenopus species form a monophyletic clade, X. tropicalis appears to have diverged from other Xenopus species prior to the formation of the extant class Ia lineages. This is not surprising because the divergence time of X. tropicalis and the Xenopus common ancestor is estimated between 50 and 81 MYA (Evans et al. 2004Go). Undoubtedly, the transspecies mode of evolution in Xenopus is strongly influenced by natural selection acting on the class Ia locus (D. H. Bos and B. Waldman, unpublished data), which tends to extend allele retention.

Pattern of Class Ia MHC Evolution in X. laevis
MHC class Ia evolution in X. laevis is more similar to MHC evolution in salmonid fishes than mammals. Salmonid fishes and Xenopus frogs show similarity in at least three aspects of MHC evolution: (1) levels of polymorphism exceeding that found in primates, (2) a distinct pattern of genetic exchange, and (3) class Ia lineages that are maintained for long periods of time. All three of these characteristics differ from the patterns of class Ia evolution found in primates. For instance, primates exhibit no allele sharing among species thought to last share a common ancestor approximately 35 MYA (Vogel et al. 1999Go), and recombination events are mostly spread throughout the gene sequence and involve very short sequence tracts (Jakobsen, Wilson, and Easteal 1998Go).

The pattern of recombination, polymorphism, and transspecies allele sharing of class Ia sequences described in salmonid fishes, and now found in X. laevis, may be the result of certain genomic features of the MHC region. Shum et al. (2001)Go suggested that these features of evolution might be due to the separation of MHC class I and class II regions onto different chromosomes. The nonlinkage of class I and class II regions may influence patterns of MHC evolution by altering the potential Hill-Robertson constraints or selection for conserved haplotype blocks and linkage disequilibrium. However, class Ia evolution in X. laevis and salmonid fishes shows similarities in polymorphism, recombination, and transspecies polymorphism. These similarities support the idea that the separation of the class I and class II regions onto different chromosomes is alone insufficient to account for these patterns of MHC evolution.

Other factors may play a role in determining class Ia gene evolution. For instance, both salmonid and X. laevis class Ia and class I–processing pathway genes are located close to one another (Namikawa et al. 1995Go; Takami et al. 1997Go; Ohta et al. 1999Go). A likely result of this linkage is that distinctive allelic associations exist among class Ia and class I–processing genes in Xenopus frogs and other taxa but are not known in primates and mice (Joly et al. 1998Go; Kaufman 1999Go; Ohta et al. 2003Go). Therefore, it is possible that the pattern of evolution common to Xenopus is due in part to the number of class Ia loci, linkage, and possible coevolution of this suite of genes. Additionally, the location of the class Ia gene in a "central" rather than "distal" position within the MHC region may influence patterns of evolution, as suggested by Nonaka et al. (1997)Go.


   Acknowledgements
TOP
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Acknowledgements
References
 
Martin Flajnik and Neil Gemmell provided useful comments on this manuscript, and Louis Du Pasquier provided ideas and invaluable technical assistance in the laboratory and in the animal breeding and care facility. Scott Edwards and two anonymous reviewers provided constructive comments on an earlier version of this manuscript. This research was supported by the Marsden Fund (Royal Society of New Zealand) and a Ph.D. scholarship from the University of Canterbury.


   Footnotes
 
1 Present address: Department of Forestry and Natural Resources, Purdue University Back

Scott Edwards, Associate Editor


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 Introduction
 Materials and Methods
 Results
 Discussion
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 References
 

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Accepted for publication August 31, 2005.


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