The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Canavez, F.
Right arrow Articles by Parham, P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Canavez, F.
Right arrow Articles by Parham, P.
The Journal of Immunology, 2001, 167: 5786-5794.
Copyright © 2001 by The American Association of Immunologists

Comparison of Chimpanzee and Human Leukocyte Ig-Like Receptor Genes Reveals Framework and Rapidly Evolving Genes1

Flavio Canavez2, Neil T. Young3, Lisbeth A. Guethlein, Raja Rajalingam, Salim I. Khakoo4, Benny P. Shum and Peter Parham5

Departments of Structural Biology and Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
The leukocyte receptor complex (LRC) on human chromosome 19 contains related Ig superfamily killer cell Ig-like receptor (KIR) and leukocyte Ig-like receptor (LIR) genes. Previously, we discovered much difference in the KIR genes between humans and chimpanzees, primate species estimated to have ~98.8% genomic sequence similarity. Here, the common chimpanzee LIR genes are identified, characterized, and compared with their human counterparts. From screening a chimpanzee splenocyte cDNA library, clones corresponding to nine different chimpanzee LIRs were isolated and sequenced. Analysis of genomic DNA from 48 unrelated chimpanzees showed 42 to have all nine LIR genes, and six animals to lack just one of the genes. In structural diversity and functional type, the chimpanzee LIRs cover the range of human LIRs. Although both species have the same number of inhibitory LIRs, humans have more activating receptors, a trend also seen for KIRs. Four chimpanzee LIRs are clearly orthologs of human LIRs. Five other chimpanzee LIRs have paralogous relationships with clusters of human LIRs and have undergone much recombination. Like the human genes, chimpanzee LIR genes appear to be organized into two duplicated blocks, each block containing two orthologous genes. This organization provides a conserved framework within which there are clusters of faster evolving genes. Human and chimpanzee KIR genes have an analogous arrangement. Whereas both KIR and LIR genes can exhibit greater interspecies differences than the genome average, within each species the LIR gene family is more conserved than the KIR gene family.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
The leukocyte receptor complex or cluster (LRC)6 occupies about 1 Mb of human chromosome 19q13.4 (1, 2, 3, 4, 5). It contains families of genes that encode receptors in which the extracellular domains are made up of Ig-like domains. One family of genes is the killer cell Ig-like receptors (KIRs) expressed by NK cells (6, 7) and subpopulations of T cells (8, 9). Adjacent to the KIR genes on their centromeric side is a distinct, but related, gene family, members of which were independently discovered by different investigators and given different names: leukocyte Ig-like receptors (LIR) (10, 11), Ig-like transcripts (ILTs) (12), and monocyte Ig-like receptors (MIRs; Table IGo) (13). Members of this second gene family encode receptors with two or four C2-type Ig domains; most are expressed by monocytes, macrophages, and dendritic cells, but some are also found on NK, B, and T cells (14, 15). In the absence of a single, agreed nomenclature, we shall use LIR here as a general descriptor for the gene family and the receptors they encode as it more accurately describes the proteins and their tissue distributions than does ILT or MIR.


View this table:
[in this window]
[in a new window]
  		Table I. Nomenclature and the GenBank accession numbers for common chimpanzee sequences1

 
Characterization of cDNA sequences combined with genomic analysis has shown the human LIR gene family consists of 11 expressed genes and two pseudogenes (4, 5). They form two homologous blocks of genes that are in opposite orientations and separated by ~200 kb (4). In accordance with the convention adopted in the human genomic analyses, 11 of the genes are named ILT1–11, the other two are named LIR6 and LIR8 (4, 5). A minor LIR gene haplotype is found lacking the expression of ILT6 (5, 16).

Within both the LIR and KIR families are subsets of receptors that have the potential for either activating or inhibitory function. The long cytoplasmic tails of inhibitory receptors contain immunoreceptor tyrosine-based inhibitory motifs (ITIMs) that interact with the tyrosine phosphatase SH2-containing phosphatase-1 to inhibit cell activation (17, 18). Activating receptors have short cytoplasmic tails that lack ITIMs, with their activating function being determined by the presence of a charged residue in the transmembrane domain. Such residues allow association with signaling adaptor molecules that possess immunoreceptor tyrosine-based activation motifs. For activating KIR the adaptor molecule is DAP-12 (19), whereas for activating LIR it is Fc{epsilon}RI{gamma} (20).

All known ligands for KIR and LIR are MHC class I or class I-like molecules (7, 14). Different KIR genes have been shown to encode receptors for HLA-A, -B, -C, and -G, and they account for approximately half of the KIR gene family (7, 14). By contrast, only LIR1 and LIR2 are known to bind MHC class I, which they do with a broad specificity, encompassing products of both classical and nonclassical HLA class I genes (17, 21, 22, 23, 24). For other members of the LIR family, the functions and ligand specificities remain unknown.

Population studies and species comparisons have shown that MHC class I genes can be highly polymorphic and rapidly evolving. Analogous studies of the KIR gene family have revealed similar characteristics. Within the human population, KIR haplotypes vary in the number and functional type of KIR genes (25, 26), and certain of the genes are also highly polymorphic (27, 28, 29). However, the LIR gene cluster appears more conserved in its organization (30). A comparison of common chimpanzee (Pan troglodytes) and human KIRs revealed that although the two species had similar numbers of KIR genes, a minority of them appeared orthologous (31). To examine the recent evolution of the LIR gene family, we have characterized common chimpanzee LIR genes and compared them to their human counterparts.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Isolation of RNA and DNA samples

A frozen spleen sample from the common chimpanzee (Pan troglodytes verus) Amanda was macerated with scalpels, strained through a 70-µm pore size cell strainer, and resuspended in RNAzol B (Tel-Test, Friendswood, TX). Total RNA was isolated from the lysate following the manufacturer’s recommended protocols. mRNA was purified using a poly(dT) cellulose column from the Poly(A) Quik mRNA Isolation kit following the manufacturer’s instructions (Stratagene, La Jolla, CA). Genomic DNA samples were prepared from 48 B lymphoblastoid cell lines of unrelated chimpanzees (31, 32, 33) following standard protocols (34). Samples of chimpanzee spleen and peripheral blood were purchased from the Yerkes Regional Primate Center at Emory University (Atlanta, GA).

cDNA library construction

A common chimpanzee spleen cDNA library was constructed with the ZAP Express Gigapack III Gold kit (Stratagene) according to the manufacturer’s instructions and screened with a human ILT2 cDNA probe following standard procedures (34). Briefly, cDNA was synthesized from 5 µg mRNA, cloned into the ZAP Express {lambda} phage vector, packaged, and used to infect the XL1-Blue strain Escherichia coli (Stratagene). For screening, 2 x 106 PFU of the primary cDNA library were plated at a density of ~1 x 105 plaques/150-mm petri dish and transferred in duplicate onto Colony/Plaque Screen nylon membranes (NEN Life Science Products, Boston, MA). The membranes were hybridized under low stringency conditions (in 30% formamide, 5x standard saline sodium phosphate-EDTA (SSPE) buffer (pH 7.4), 5x Denhardt’s reagent, 5% dextran sulfate, 1% SDS, and 1 mg/ml fragmented and denatured salmon sperm DNA at 42oC for 16 h) with a human ILT2 cDNA probe, washed four times with 2x SSPE/0.5% SDS at 42oC, and autoradiographed. Positive phage were plaque-purified and subjected to an in vivo excision procedure according to a Stratagene protocol; the resulting phagemids contained chimpanzee cDNA within the pBK-CMV vector.

Nucleotide sequencing and alignment

Nucleotide sequences were determined on both DNA strands using the Big-Dye Terminator Cycle Sequencing kit (Applied Biosystems, Foster City, CA) and an ABI 377 DNA sequencer (Applied Biosystems). Complete DNA sequences of the plasmid inserts were obtained by using T3 and T7 universal oligonucleotide primers and internal primers based on sequences of chimpanzee LIRs. DNA sequences were assembled and analyzed using the computer program AutoAssembler (version 2.1; Applied Biosystems) and the Wisconsin Package sequence analysis software (version 10.1; Genetics Computer Group, Madison, WI) (35).

LIR gene nomenclature

Individual cDNA sequences in the human LIR family have been named, mainly using two different nomenclatures: either ILT (12) or LIR (10, 11), followed by a number in series (Table IGo). These alternative and competing nomenclatures have yet to be reconciled into a single system. In human genomic analyses a hybrid nomenclature is being used (4, 5) in which the preference is for the ILT nomenclature (11 of the genes are named ILT1–11), but LIR nomenclature is retained for two genes (LIR6 and LIR8, first described by Cosman et al., who introduced the LIR nomenclature) (10, 11). Common chimpanzee LIR genes that are clearly orthologous to a human LIR gene have been given the same names as the human genes with the additional prefix Pt- for Pan troglodytes. Chimpanzee LIR genes that are paralogous to human LIR genes or for which orthology is less certain have been given provisional designations using lowercase letters: Pt-LIRa-e. The chimpanzee cDNA sequences reported here were deposited into the GenBank database (Table IGo).

5' RACE

We performed 5' RACE analysis to obtain the 5' coding region sequences of certain incomplete chimpanzee LIR cDNAs. First-strand cDNA was synthesized using 1 µg mRNA isolated from the chimpanzee Amanda with the SMART RACE kit according to the manufacturer’s instructions (Clontech, Palo Alto, CA). Two specific reverse oligonucleotide primers were designed for each chimpanzee LIR gene based on sequences determined for partial cDNAs isolated from library screening. These oligonucleotide primers were used for amplifying the 5' cDNA of desired genes in two rounds of PCR amplifications. The PCR conditions were 94oC for 30 s; 25 cycles of 94oC for 5 s, 66oC for 10 s, and 72oC for 3 min; and 72oC for 3 min for the initial reactions and 94oC for 30 s; 15 cycles of 94oC for 5 s, 65oC for 10 s, 72oC for 3 min; and 72oC for 3 min for the second-round nested amplifications. Antisense oligonucleotide primers used in the initial amplification were 5'-GGG CTC AGA TCA CAG GAC TCA CG-3' (Pt-LIRc), 5'-CGG GCA TGG GAA TGG GAG TTC AGA C-3' (Pt-LIRd), and 5'-CTG TCG GTC AGG GCG CTG GGC G-3' (Pt-ILT7). Antisense primers used in the second-round nested PCR were 5'-CCA CAC CTG GTC GTT GTA AGT AT-3' (Pt-LIRc), 5'-CTT GCG TGT TCC CAG GTG ATG GAC G-3' (Pt-LIRd), and 5'-GTC AGA TTC TCT CCG GGG GTC ACA A-3' (Pt-ILT7). Amplified cDNA fragments were separated by electrophoresis in 1% agarose gel, purified using the QIAEX II gel extraction kit (Qiagen, Chatsworth, CA), cloned into the pCR-TOPO plasmid vector (Invitrogen, Carlsbad, CA), and sequenced.

Phylogenetic and recombination analyses

Phylogenetic analyses were performed separately with nucleotide sequences encoding for the entire mature proteins and for only the Ig-like domains, using the PAUP 4.0 (Phylogenetic Analysis Using Parsimony) software package (version 4.0b4a, Sinauer Associates, Sutherland, MA) and other methods. Phylogenetic trees were constructed using the neighbor-joining method (36), and confidence values for each tree branch were estimated from 1000 replicates using the bootstrap technique (37). Gaps were counted as single mutational events in pairwise comparisons. Initial analysis to assess recombination between LIR genes was performed using the Partimatrix program (38), including all chimpanzee and human LIR gene sequences. Further analyses were made with groups of four sequences (two each from chimpanzee and human) and also with clusters of LIR sequences identified from phylogenetic analyses.

Analysis of nonsynonymous and synonymous nucleotide substitutions

Rates of dN (nonsynonymous) and dS (synonymous) substitutions were determined using the synonymous/non-synonymous analysis program (SNAP, available at http://hiv-web.lanl.gov/SNAP/WEBSNAP/SNAP.html) based on the method of Nei and Gojobori (39) and incorporating a statistic developed by Ota and Nei (40). This analysis was performed separately on complete LIR mature protein-coding sequences, on sequences encoding all Ig-like domains, on individual Ig-like domains, or on contiguous small regions (60 nt) of each Ig-like domain. Each chimpanzee LIR gene was compared with the most closely related human LIR sequence.

Genomic typing for common chimpanzee LIR genes

To type for the presence of LIR genes in a panel of 48 unrelated chimpanzees, we designed two pairs of gene-specific PCR oligonucleotide primers for each of the nine chimpanzee LIRs that anneal to exon sequences encoding the first and second Ig-like domains (Table IIGo). PCRs were conducted with ~100–200 ng genomic DNA, 25 pmol of each primer, 2.5 mM of each dNTP, 1.5 mM MgCl2, 1x AmpliTaq buffer (Applied Biosystems), and 1 U AmpliTaq DNA polymerase (Applied Biosystems). The PCR parameters were 94oC for 10 s; 30 cycles of 94oC for 40 s, 68oC for 30 s, and 72oC for 30 s; and 72oC for 10 min. Amplified genomic DNA fragments were analyzed on ethidium bromide-stained 1% agarose gels. Typing reactions were tested on genomic DNA of the common chimpanzee Amanda from whom the spleen cDNA library was constructed; the PCR-amplified DNA were directly sequenced to confirm specificities. Subsequently, genomic DNA from a panel of 48 unrelated chimpanzees were typed, including individuals from three common chimpanzee subspecies: P. troglodytes schweinfurthii, P. troglodytes troglodytes, and P. troglodytes verus.


View this table:
[in this window]
[in a new window]
  		Table II. Oligonucleotide primers used for typing chimpanzee genomic DNA1

 

   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Isolation and characterization of cDNAs encoding nine chimpanzee LIR genes

A cDNA library was made from spleen cells of an individual chimpanzee and screened with a human ILT2 cDNA probe. The screen yielded 83 hybridizing cDNA clones, which were further characterized by DNA sequencing using the T3 and T7 phagemid-based oligonucleotide primers. On the basis of partial sequences in the 5' and 3' ends of the cDNAs, the clones were sorted into nine groups, of which four were well represented. The other five groups were represented by only one clone each, three of these being incomplete clones lacking the 5' end. Of the 83 clones 41 appeared to be full-length cDNA clones; the remaining 42 clones appeared either as partial clones lacking 5' regions or products of alternatively spliced variants. A total of 44 cDNA clones representing all nine groups were completely sequenced. For the three groups represented by only single partial clones, complete coding region sequences were characterized using 5' RACE analysis. All nine sequences were novel, but clearly belong to the gene family defined by the human LIR sequences.

Comparison of chimpanzee and human LIR

Eight of the nine chimpanzee LIR genes encode proteins with four extracellular Ig-like domains (D1–D4); the ninth encodes a protein with two Ig-like domains. This strong bias toward receptors with four Ig-like domains parallels that seen in the human LIR family (14). The positions of cysteine residues that make up the intradomain disulfide bonds of the Ig-like domains are conserved in chimpanzee and human. Aligning and comparing the chimpanzee and human LIR as a group reveals sequence variability of ~78% in D1, ~72% in D2, ~49% in D3, and ~64% in D4. Variability is well spread within all Ig-like domains, although D3 appears to the most conserved (data not shown).

To identify affinities between individual chimpanzee and human LIR, we performed a systematic pairwise comparison of amino acid sequences. Four of the chimpanzee sequences had particular affinity with one human sequence, consistent with them being pairs of orthologous genes (Fig. 1Go and identified by arrows). Within each of the four orthologous pairs, the amino acid sequences were <5% different, whereas differences between the four chimpanzee orthologs and other human paralogs were >17%. The human LIR shown to have chimpanzee orthologs were ILT1, ILT3, ILT7, and LIR8; we therefore gave these chimpanzee LIR the same name as their human orthologs and also the prefix Pt- for Pan troglodytes. For the other five chimpanzee LIRs, the patterns of pairwise difference are distinct from those seen for the four with obvious human orthologs (Fig. 1Go). There is a more continuous range of differences, in which no single human LIR is as closely related as seen with the orthologous pairs. To indicate this difference, these chimpanzee LIRs have been given names that do not correspond to human LIRs: Pt-LIRa, -b, -c, -d, and -e. These five chimpanzee LIRs can be further divided into two groups according to the groups of human LIR to which they are more closely related. Thus, Pt-LIRa, -b, and -c are more closely related to human ILT1, -2, -4, and -6 and LIR6, whereas Pt-LIRd and -e are closer to human ILT5, -8, and -9 (Figs. 1Go and 2Go). These affinities could reflect orthologous or paralogous relationships. None of the chimpanzee LIRs shows particular affinity with ILT11 or the pseudogene ILT10 (Fig. 2Go). Within the population of cDNA clones we sequenced, the relative abundance of chimpanzee LIRs were: 32% for Pt-LIRa, 32% for Pt-LIRb, 14% for Pt-LIR8, 12% for Pt-ILT1, and 2% each for Pt-ILT3, ILT7, LIRc, LIRd, and LIRe.



View larger version (40K):
[in this window]
[in a new window]
  		FIGURE 1. Pairwise comparison of amino acid sequences of common chimpanzee and human LIRs identifies four pairs of orthologs. Pairwise differences were calculated from amino acid sequences of the mature proteins and were plotted as percentages. Vertical arrows mark comparisons of orthologs.

 


View larger version (34K):
[in this window]
[in a new window]
  		FIGURE 2. A schematic comparison of the proteins encoded by 9 chimpanzee and 13 human LIR genes. Orthologs are depicted on the same line. Protein domains are shown as rectangles, with D1, D2, D3, and D4 corresponding to the Ig-like domains and S, T, and CY corresponding to the stem, transmembrane, and cytoplasmic regions, respectively. +, a conserved arginine residue in the transmembrane domain of short-tailed LIRs. {diamond} and {diamondsuit}, Positions of ITIMs; {diamond}, YxxV motif; {diamondsuit}, YxxL motif. The pseudoexon in the region encoding the cytoplasmic tail of Pt-ILT3 is represented by a solid horizontal line. Dashed lines in the homologous regions of human ILT4 and LIR8 indicate unknown data. A, The three orthologous pairs that are more divergent from the other LIRs (see also Fig. 3Go). B, The orthologous Pt-ILT1/ILT1 and related LIRs that form a separate cluster (I) in Fig. 3Go. C, Another group of paralogous chimpanzee and human LIRs, including the pseudogene ILT9, that form a distinguishable cluster (II) in Fig. 3Go. D, The two divergent human LIR (ILT11 and the pseudogene ILT10) that are not represented in the chimpanzee LIRs here defined. Putative domain designation of the chimpanzee LIRs, human ILT11, and pseudogenes ILT9 and ILT10 were obtained from comparison with other human LIRs.

 
The similarities and differences in the chimpanzee and human LIR proteins are depicted in the schematic comparison of Fig. 2Go. The orthologous LIRs include two with four Ig-like domains and short cytoplasmic tails (Pt-ILT1, Pt-ILT7), one with four Ig-like domains and a long cytoplasmic tail (Pt-LIR8), and one with two Ig-like domains and a long cytoplasmic tail (Pt-ILT3). Each group of paralogous LIR contains members with long and short cytoplasmic tails (Fig. 2Go, B and C). All short-tailed chimpanzee LIR contains an arginine residue in the transmembrane region homologous to those found in human short-tailed LIRs and which is implicated in association with an activating adaptor molecule. The three or four ITIMs present in the long-tailed chimpanzee LIR could be important for inhibitory function. The chimpanzee LIRs with three ITIMs (Pt-ILT3, Pt-LIR8, Pt-LIRb) lack either the first or the second ITIM compared with human and chimpanzee LIRs that have four ITIMs. With one exception, the first pair of ITIMs has YxxV motifs, and the second pair has YxxL motifs; the exception is the last ITIM of the chimpanzee Pt-ILT3, which has a YxxV motif. Pt-ILT3 also has a deletion in the cytoplasmic tail that is not present in its human ortholog, but is identical with that present in human ILT4 and LIR8. Sequence analysis by PCR amplification of this region of the Pt-ILT3 gene showed the presence of a pseudoexon that encodes the missing part of the cytoplasmic tail (data not shown). That the pseudoexon is not incorporated into mRNA was confirmed by RT-PCR analysis of Pt-ILT3 from several individual chimpanzees (data not shown).

Phylogenetic trees were constructed from the human and chimpanzee LIR nucleotide sequences. Shown in Fig. 3Go are two trees, one made from nucleotide sequences encoding the mature proteins, and the other made only from sequences encoding the extracellular Ig-like domains. Both trees have a similar topology consisting of five main clusters of LIR, each containing both chimpanzee and human LIR, and two additional branches represented by single human sequences (ILT10 and ILT11). Comparison of the amino acid sequences also gave trees with similar topology (data not shown). Three of the clusters (III, IV, and V in Fig. 3Go) contain only a pair of orthologous LIRs: ILT3/Pt-ILT3, LIR8/Pt-LIR8, and ILT7/Pt-ILT7. Cluster I in Fig. 3Go involves four chimpanzee and five human LIRs, including the orthologs ILT1/Pt-ILT1. Cluster II of two chimpanzee and three human sequences contains no orthologous LIRs. In both trees Pt-LIRb and ILT4 (LIR2) form a separate branch, raising the possibility that they are orthologs. However, they are more diverged from each other than is the case for the other orthologous pairs, and in both trees the confidence in the ILT4/Pt-LIRb branch (86 and 83%) is considerably less than for ILT1/Pt-ILT1, ILT3/Pt-ILT3, ILT7/Pt-ILT7, and LIR8/Pt-LIR8 (100%).



View larger version (44K):
[in this window]
[in a new window]
  		FIGURE 3. Phylogenetic analyses of chimpanzee and human LIR. Consensus trees were obtained using the neighbor-joining method (36 ) with the Kimura two-parameter distance correction (46 ). The tree in A was generated from nucleotide sequences encoding the mature protein, and the tree in B was generated from sequences encoding only the Ig-like domains. Numbers on nodes are percent bootstrap values calculated from 1000 replications using the heuristic search parameters of the PAUP program (Sinauer Associates). Clusters shaded in a darker gray are major groupings containing both human and chimpanzee sequences and are represented in both trees; these clusters are also noted by Roman numerals. The lighter gray shading indicates potential orthologs found within cluster I.

 
To assess the role of recombination in generating LIR gene diversity, we used the Partimatrix program described by Jakobsen et al. (38). The extent to which individual LIR genes have been involved in recombination varies. Those genes for which human/chimpanzee orthologies were readily identified (Fig. 1Go) and which were defined with deeper branches and high confidence in the phylogenetic analyses (Fig. 3Go) have not been involved in recombination: these comprise ILT3/Pt-ILT3, ILT7/Pt-ILT7, and LIR8/Pt-LIR8. An example of output given for such genes by the Partimatrix program is shown for ILT3/Pt-ILT3 in Fig. 4GoA. It shows that there has been little recombination between genes in two major clusters of the phylogenetic trees (ILT5/Pt-LIRd vs ILT3/Pt-ILT3; Figs. 3Go and 4GoA).



View larger version (44K):
[in this window]
[in a new window]
  		FIGURE 4. Recombination analyses of chimpanzee and human LIR. Outputs from the program Partimatrix (38 ) are shown. Shown above each matrix are the partitions into which the data were grouped (each column of circles represents an individual partition); filled circles indicate the group being considered for each partition. Each row of boxes corresponds to an informative nucleotide position along the cDNAs; the protein domains to which they contribute are noted with L for the leader peptide, D1–4 for Ig-like domains 1–4, S for the stem region, T for transmembrane domain, and CY for the cytoplasmic domain. Open boxes show partitions in agreement at the given nucleotide position; furthermore, a dot in the box indicates perfect agreement with the partition, and a slash indicates positions for which two or more partitions are in perfect agreement. Filled boxes show positions that disagree with the partition shown at the head of the column. A, An example for which no recombination was detected ,as can be seen by the fact that only two sites disagree with grouping of ILT3 and Pt-ILT3. B, Potential recombinations within the group comprised of Pt-LIRd, Pt-LIRe, ILT5, ILT8, and ILT9 (cluster II in Fig. 3Go). C, More extensive recombination within the group comprised of Pt-LIRa, Pt-LIRb, Pt-LIRc, ILT2, ILT4, and ILT6 (from cluster I in Fig. 3Go).

 
Partition matrix analysis of all five LIRs in cluster II (Fig. 3Go), the cluster containing no obviously orthologous genes, shows that recombination has played a considerable role in the diversification of these five LIRs (Fig. 4GoB). For example, ILT5 and ILT8 are very similar in the region encoding the extracellular part of the molecule (column 3 in Fig. 4GoB), but they diverge at the 3' exons encoding the transmembrane and cytoplasmic domains. Here, ILT5 groups with Pt-LIRd (column 2 in Fig. 4GoB), and ILT8 with Pt-LIRe (column 1 in Fig. 4GoB), reflecting the divergence of sequences encoding the long-tail and short-tail LIRs. That recombination between exons encoding the Ig-like domains and the 3' exons has been instrumental in diversifying cluster II LIRs is further supported by differences in the fine structures of this cluster in phylogenetic trees made from sequences encoding mature proteins (Fig. 3GoA) and only exons encoding the Ig-like domains (Fig. 3GoB). A more complicated pattern of recombination is revealed in the Partimatrix analysis of six members from cluster I (Fig. 3Go): ILT2, -4, and -6 and Pt-LIRa, -b, and -c (Fig. 4GoC).

Each chimpanzee LIR was compared with the most closely related human LIR(s), and the relative frequencies of nonsynonymous (dN, coding) and synonymous (dS, silent) substitutions were calculated. Overall comparison of the sequences encoding the mature protein revealed an excess of synonymous substitutions for all LIRs, consistent with the action of purifying selection upon LIR genes (Table IIIGo). When sequences encoding only the Ig-like domains were compared, some ratios were higher and closer to 1.00, the value expected under neutral evolution. For the Pt-LIRa/LIR6 and Pt-LIR8/LIR8 comparisons the values were in slight excess of 1.00 (Table IIIGo). To refine the analysis we divided the sequence of each Ig-like domain into five similarly sized fragments of ~60 nt and computed the dN and dS values for these fragments. The values are consistent with purifying selection for almost all regions of the Ig-like domains of ILT1/Pt-ILT1, ILT3/Pt-ILT3, and ILT7/Pt-ILT7, with a few isolated regions containing only nonsynonymous substitutions (Fig. 5GoA). In contrast, the values for LIR8/Pt-LIR8 provide evidence for positive selection throughout the fourth Ig-like domain and in the carboxyl-terminal part of the first and second Ig-like domains (Fig. 5GoA). Comparisons of Pt-LIRd and Pt-LIRe with human ILT5, -8, and -9 gave no clear evidence for positive selection overall; however dN was locally larger than dS in segments 10 and 20 of the Pt-LIRd analysis and segment 9 of the Pt-LIRe analysis (Fig. 5GoB). Comparison of Pt-LIRa, Pt-LIRb, and Pt-LIRc with human ILT2, ILT4, and LIR6 also revealed no evidence for positive selection (data not shown).


View this table:
[in this window]
[in a new window]
  		Table III. dN:dS ratios estimated for human and chimpanzee LIR genes1

 


View larger version (40K):
[in this window]
[in a new window]
  		FIGURE 5. Comparison of the distribution of synonymous (dS) and nonsynonymous (dN) substitutions in chimpanzee and human LIR sequences. Each Ig-like domain-coding region was divided into small fragments of ~60 nt each. Analyses were made for each of these fragments and are plotted as a histogram. {square}, dS values; {blacksquare}, dN. D1-, D2-, D3-, and D4-coding regions comprise fragments 1–5, 6–10, 11–15, and 16–20, respectively. A, Analysis of the four orthologous chimpanzee and human LIRs. B, comparison of the unassigned chimpanzee genes, Pt-LIRd and Pt-LIRe, when compared with the three closest human LIRs, i.e., ILT5, ILT8, and ILT9. In these analyses, each fragment is represented by six bars (three dS and three dN): the two bars on the left show results generated by analyses with ILT5, the two bars in the middle show results generated by analyses with ILT8, and the two bars on the right show results generated by analyses with ILT9.

 
Within species LIR haplotypes are more conserved than KIR haplotypes

A characteristic of chimpanzee and human KIR haplotypes is variation in the number and content of genes. To determine whether this is also a feature of the chimpanzee LIR gene family, we developed a PCR-based system for typing the nine chimpanzee LIRs and used it to type genomic DNA from a panel of 48 common chimpanzees that had already been typed for Pt-KIR (31). Whereas 30 different Pt-KIR genotypes are represented in this panel, only two different LIR genotypes were identified (Fig. 6Go). The common genotype consisted of all nine Pt-LIRs and was observed in 42 individuals representing two subspecies, P. troglodytes schweinfurthii and P. troglodytes verus. The minority genotype was defined by negative typing for Pt-LIRa and was seen in six individuals representing three subspecies: schweinfurthii, troglodytes, and verus. Thus there is evidence for some heterogeneity in Pt-LIR haplotypes, but it is modest compared with the variability seen with Pt-KIRs.



View larger version (73K):
[in this window]
[in a new window]
  		FIGURE 6. SSP-PCR typing of LIR and KIR genes from the genomic DNA of 48 unrelated common chimpanzees. Common chimpanzee LIR genes are divided into those with long, short, or no (for potentially soluble molecules) cytoplasmic tails, while KIRs are divided into those with long and short tails. LIR typings are shown in dark gray, and KIR typings are shown in light gray. The frequency of each gene in the panel is shown at the bottom.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
LIR and KIR are related gene families closely linked in the human genome, and both encode activating and inhibitory leukocyte receptors in which the extracellular regions are made up of Ig-like domains. Neither of these gene families appears to be present in the mouse genome, at least not as a recognizable ortholog, suggesting that these receptor systems have undergone considerable change and diversification during the course of mammalian evolution (41). Investigation of this proposition requires comparison of species more closely related than mice and men; therefore, we have been studying chimpanzees, the living species most closely related to humans. This study aimed to identify the expressed LIR genes of the common chimpanzee (Pan troglodytes) and to determine their structural and evolutionary relationships with human LIRs.

Thirteen human LIR genes (ILT1–11, LIR6, and LIR8) have been defined, of which two appear to be pseudogenes (ILT9 and ILT10) (4, 5). From the analysis of cDNA clones obtained from a splenocyte library we have characterized nine different chimpanzee LIRs, all expressed in the cells of one individual. The nine chimpanzee and 11 human expressed LIRs exhibit a similar range of diversity, as is clearly seen in phylogenetic trees (Fig. 3Go). In these trees are five LIR clusters, of which two contain only long-tailed LIRs (III and IV), one contains only short-tailed LIRs (V), and two contain both long- and short-tailed LIRs (I and II). All these clusters and subclusters contain both chimpanzee and human LIR. Thus, the set of nine chimpanzee LIRs defined here should provide good representation of the total set of chimpanzee LIR genes, although the possibility that additional family members await discovery must not be ruled out.

Genotyping 48 unrelated common chimpanzees showed 42 to have genes for all nine chimpanzee LIRs; the remaining six individuals had just eight of the genes. The negative typing reactions for Pt-LIRa could be due to absence of the gene or to Pt-LIRa gene polymorphism that affected the oligonucleotide priming sites used in the typing. Favoring the former interpretation is the fact that two different sets of Pt-LIRa-specific oligonucleotide primers gave negative results. Either way, there is clear evidence for two different chimpanzee LIR haplotypes with frequencies of 35% (Pt-LIRa-) and 65% (Pt-LIRa+) assuming Hardy-Weinberg equilibrium. Two human LIR haplotypes have also been defined, one containing all the genes and one containing an ILT6 psuedogene with a ~6.7-kb deletion (5, 16); the latter haplotype appeared to be have a frequency of 18–28% in the populations examined (5, 16). The persistence of such deletion haplotypes is evident by their presence in common chimpanzee subspecies that diverged about 1.6 million years ago (42). The modest variation in LIR genotypes observed within chimpanzee and human contrasts dramatically with the extensive variation in the KIR genotype observed in both species (Fig. 6Go) (25, 31).

Pairwise comparison of chimpanzee and human LIR allowed four pairs of orthologs to be assigned with confidence (Fig. 1Go): ILT1/Pt-ILT1, ILT3/Pt-ILT3, ILT7/Pt-ILT7, and LIR8/Pt-LIR8. These pairs of nucleotide sequences differ by about 2%, comparable to estimates of the overall genome sequence similarity of ~98.8% between human and chimpanzee (43). These genes show little evidence of having undergone recombination with other LIR genes, and in large part the differences between the species do not appear to be the result of natural selection; exceptional in this regard are LIR8 and Pt-LIR8 (Fig. 5GoA). On the basis of cDNA sequence comparison, orthologous relationships cannot be assigned between five chimpanzee and nine human LIRs. In the pairwise comparisons of these five chimpanzee LIR sequences, they differ by 9–14% in nucleotides with the closest human LIRs, values much greater than the genome average and which appear largely to be a consequence of recombination between LIR genes. Attempts to identify chimpanzee genes more closely related to human ILT2, -4, -5, -8, and -11 were made by PCR amplification of chimpanzee genomic DNA with primers specific for these human genes. No positive reaction was obtained (data not shown) consistent with the absence of conserved, orthologs for these genes.

Comparison of chimpanzee and human LIR allows us to divide the LIR gene family into a group of four genes that have been relatively stable during the ~5.5 million yr of evolution that separate modern chimpanzees and humans (44), and an additional group of genes that have been evolving more rapidly through intergenic recombination, gene duplication, and gene deletion. In the human LRC, LIR genes are organized as two similarly sized regions (4). These arose by en bloc duplication of an ancestral region, which gave rise to two daughter blocks in opposite transcriptional orientations and separated by an intervening region of about 200 kb (4, 16). We now see that the four human LIR genes having chimpanzee orthologs are evenly distributed between the two blocks, and each block contains one long-tailed and one short-tailed LIR (LIR8 and ILT7 in one block, and ILT1 and ILT3 in the other; Fig. 7Go).



View larger version (24K):
[in this window]
[in a new window]
  		FIGURE 7. A schematic representation of the LIR gene cluster in human and common chimpanzee. The upper portion of the figure shows the genomic organization of the human LIR (4 5 ). The arrows indicate the direction of transcription for each gene. Shown below is a model for the organization of the chimpanzee LIR locus as deduced from phylogenetic analysis. Orthologous genes are shown as the taller rectangles. Shading is based on the ancestral relationships previously proposed (4 ), which are in agreement with the analyses performed here. One ancestral group described by Wende et al. (4 ) corresponds to cluster I, and the other ancestral group to clusters II–IV. Finally, cluster V (ILT7/Pt-ILT7) was proposed to be a translocation/duplication that occurred subsequent to the initial inverse duplication.

 
In humans, two clusters of rapidly evolving genes in each duplicated block are separated from each other by the orthologous or framework genes (Fig. 7Go). Regarding the nonorthologous genes, ILT4 and related members (cluster I) lie between LIR8 and ILT7 in one block, and between ILT1 and ILT3 in the other. In contrast, ILT5 and related genes (cluster II) are downstream of LIR8 and ILT3. Thus the organization of the LIR regions is such that conserved, orthologous framework genes alternate with more rapidly evolving genes. This situation is like that found in the KIR gene family where three framework genes define two intervals of high gene variability (5, 31, 45). Also comparable in the two gene families are the percent sequence identities between human and chimpanzee orthologs (~2–5%) and more rapidly evolving genes (>8%). Furthermore, the numbers of inhibitory, long-tailed KIR and LIR are similar in humans and chimpanzees, but humans have more activating, short-tailed KIR and LIR than chimpanzees. Thus, for both families the inhibitory receptors appear more conserved.

Fig. 7Go shows a working model for the organization of the chimpanzee LIR complex and its comparison to human LIR genes. Given the good representation of human and chimpanzee LIR genes in all major clusters, the chimpanzee LIR gene family is probably organized similarly to the human LIR genes in two homologous blocks (Fig. 7Go). Chimpanzee genes have been placed in the same intervals as human genes. In this scheme one can have some confidence in the positions of the orthologous or framework genes, but the positions assigned to the faster evolving clusters of ILT4- and ILT5-related genes chimpanzee genes remain speculative. Consider first the cluster consisting of ILT4 and related genes (cluster I in Fig. 3Go). As Pt-LIRb groups with human ILT4 in both phylogenetic analyses (Fig. 3Go), and since they both encode long-tailed LIRs, we assigned them to the same interval. As Pt-LIRc is related to human ILT6 (Fig. 3BGo), and as they are the only genes encoding soluble proteins (Fig. 2Go), Pt-LIRc could be assigned to the same interval as ILT6. Alternatively, ILT6 and Pt-LIRa could be placed at the same interval, as they are both the missing genes on the minority haplotypes. Of the human genes in the ILT5-related cluster (II in Fig. 3Go), ILT5 and ILT8 are together in one gene block and represent a closely related pair encoding an inhibitory and an activating four-Ig-domain receptor. In contrast, the ILT5-related gene of the other block is ILT9, an unusual LIR with three Ig-like domains. As Pt-LIRd and Pt-LIRe encode a conventional pair of inhibitory and activating LIR, such as ILT5 and ILT8, these genes have been assigned to the same interval as ILT5 and ILT8, respectively. These tentative assignments provide a working model for future direct analysis of the genomic organization of the chimpanzee LIR gene family.

In both humans and chimpanzees, their KIR haplotypes are much more diverse in gene number and content than are the LIR haplotypes. The propensity for new KIR haplotypes to evolve by asymmetric recombination is explained by the close proximity of the KIR genes and their separation by short, highly homologous intergenic sequences (5). This is not the situation for the human LIR gene family (4, 5). Due to this difference we anticipated that the LIR genes would be phylogenetically more conserved than the KIR genes. This assumption was proven wrong, with the interspecies variation in KIR and LIR genes being comparable. Both families contain some genes that are relatively stable and recognizable as orthologs, whereas other genes have been rapidly evolving through recombination. Thus, for some genes in both the KIR and LIR families there appears to be natural selection for new variants. The KIR genes known to encode receptors with specificity for classical MHC class I molecules are ones that have rapidly evolved since divergence of human and chimpanzee ancestors (31). Similarly, the LIRs with known specificity for MHC class I are encoded by genes, ILT2 (LIR1) and ILT4 (LIR2), with no clear-cut chimpanzee orthologs and that are members of a cluster of rapidly evolving genes. In conclusion, this comparison of chimpanzee and human species shows that both the LIR and KIR gene families evolve rapidly, making the LRC a likely hotspot of difference between chimpanzee and human genomes.


   Acknowledgments
 
We thank the Yerkes Regional Primate Center at Emory University for providing chimpanzee samples. B.P.S. thanks particularly the staff at Yerkes for their hospitality and for their assistance in obtaining chimpanzee spleen samples. We also thank Dr. Erin Adams for the generation of the chimpanzee B cell lines.


   Footnotes
 
1 This work was supported by the National Institutes of Health Grant AI31168 (to P.P.). S.I.K. was a fellow of the Cancer Research Institute (New York). N.T.Y. was a Wellcome Trust International Prize Travelling Research Fellow (United Kingdom). Back

2 Current address: Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo SP 05508-900, Brazil. Back

3 Current address: Nuffield Department of Surgery, University of Oxford, John Radcliffe Hospital, Oxford, U.K. OX3 9DU. Back

4 Current address: Department of Cell and Molecular Medicine, Southampton General Hospital, Tremona Road, Southampton, U.K. S016 6YD. Back

5 Address correspondence and reprint requests to Dr. Peter Parham, Department of Structural Biology, Stanford University School of Medicine, 299 Campus Drive West, Sherman Fairchild Building, D-151, Stanford, CA 94305-5126. E-mail address: peropa{at}stanford.edu Back

6 Abbreviations used in this paper: LRC, leukocyte receptor complex or cluster; ILT, Ig-like transcript; ITIM, immunoreceptor tyrosine-based inhibition motif; KIR, killer cell Ig-like receptor; LIR, leukocyte Ig-like receptor; MIR, monocyte Ig-like receptor; SSP, sequence-specific primers; SSPE, standard saline sodium phosphate-EDTA buffer. Back

Received for publication July 19, 2001. Accepted for publication September 6, 2001.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Torkar, M., Z. Norgate, M. Colonna, J. Trowsdale, M. J. Wilson. 1998. Isotypic variation of novel immunoglobulin-like transcript/killer cell inhibitory receptor loci in the leukocyte receptor complex. Eur. J. Immunol. 28:3959.[Medline]
  2. Wende, H., M. Colonna, A. Ziegler, A. Volz. 1999. Organization of the leukocyte receptor cluster (LRC) on human chromosome 19q13.4. Mamm. Genome 10:154.[Medline]
  3. Liu, W. R., J. Kim, C. Nwankwo, L. K. Ashworth, J. P. Arm. 2000. Genomic organization of the human leukocyte immunoglobulin-like receptors within the leukocyte receptor complex on chromosome 19q13.4. Immunogenetics 51:659.[Medline]
  4. Wende, H., A. Volz, A. Ziegler. 2000. Extensive gene duplications and a large inversion characterize the human leukocyte receptor cluster. Immunogenetics 51:703.[Medline]
  5. Wilson, M. J., M. Torkar, A. Haude, S. Milne, T. Jones, D. Sheer, S. Beck, J. Trowsdale. 2000. Plasticity in the organization and sequences of human KIR/ILT gene families. Proc. Natl. Acad. Sci. USA 97:4778.[Abstract/Free Full Text]
  6. Lanier, L. L.. 1998. NK cell receptors. Annu. Rev. Immunol. 16:359.[Medline]
  7. Long, E. O., S. Rajagopalan. 2000. HLA class I recognition by killer cell Ig-like receptors. Semin. Immunol. 12:101.[Medline]
  8. D’Andrea, A., L. L. Lanier. 1998. Killer cell inhibitory receptor expression by T cells. Curr. Top. Microbiol. Immunol. 230:25.[Medline]
  9. Uhrberg, M., N. M. Valiante, N. T. Young, L. L. Lanier, J. H. Phillips, P. Parham. 2001. The repertoire of killer cell Ig-like receptor and CD94:NKG2A receptors in T cells: clones sharing identical {alpha}{beta} TCR rearrangement express highly diverse killer cell Ig-like receptor patterns. J. Immunol. 166:3923.[Abstract/Free Full Text]
  10. Cosman, D., N. Fanger, L. Borges, M. Kubin, W. Chin, L. Peterson, M. L. Hsu. 1997. A novel immunoglobulin superfamily receptor for cellular and viral MHC class I molecules. Immunity 7:273.[Medline]
  11. Borges, L., M. L. Hsu, N. Fanger, M. Kubin, D. Cosman. 1997. A family of human lymphoid and myeloid Ig-like receptors, some of which bind to MHC class I molecules. J. Immunol. 159:5192.[Abstract]
  12. Samaridis, J., M. Colonna. 1997. Cloning of novel immunoglobulin superfamily receptors expressed on human myeloid and lymphoid cells: structural evidence for new stimulatory and inhibitory pathways. Eur. J. Immunol. 27:660.[Medline]
  13. Wagtmann, N., S. Rojo, E. Eichler, H. Mohrenweiser, E. O. Long. 1997. A new human gene complex encoding the killer cell inhibitory receptors and related monocyte/macrophage receptors. Curr. Biol. 7:615.[Medline]
  14. Allan, D. S., A. J. McMichael, V. M. Braud. 2000. The ILT family of leukocyte receptors. Immunobiology 202:34.[Medline]
  15. Colonna, M., H. Nakajima, M. Cella. 2000. A family of inhibitory and activating Ig-like receptors that modulate function of lymphoid and myeloid cells. Semin. Immunol. 12:121.[Medline]
  16. Torkar, M., A. Haude, S. Milne, S. Beck, J. Trowsdale, M. J. Wilson. 2000. Arrangement of the ILT gene cluster: a common null allele of the ILT6 gene results from a 6.7-kbp deletion. Eur. J. Immunol. 30:3655.[Medline]
  17. Fanger, N. A., D. Cosman, L. Peterson, S. C. Braddy, C. R. Maliszewski, L. Borges. 1998. The MHC class I binding proteins LIR-1 and LIR-2 inhibit Fc receptor-mediated signaling in monocytes. Eur. J. Immunol. 28:3423.[Medline]
  18. Burshtyn, D. N., A. S. Lam, M. Weston, N. Gupta, P. A. Warmerdam, E. O. Long. 1999. Conserved residues amino-terminal of cytoplasmic tyrosines contribute to the SHP-1-mediated inhibitory function of killer cell Ig-like receptors. J. Immunol. 162:897.[Abstract/Free Full Text]
  19. Lanier, L. L., B. C. Corliss, J. Wu, C. Leong, J. H. Phillips. 1998. Immunoreceptor DAP12 bearing a tyrosine-based activation motif is involved in activating NK cells. Nature 391:703.[Medline]
  20. Nakajima, H., J. Samaridis, L. Angman, M. Colonna. 1999. Human myeloid cells express an activating ILT receptor (ILT1) that associates with Fc receptor {gamma}-chain. J. Immunol. 162:5.[Abstract/Free Full Text]
  21. Colonna, M., F. Navarro, T. Bellon, M. Llano, P. Garcia, J. Samaridis, L. Angman, M. Cella, M. Lopez-Botet. 1997. A common inhibitory receptor for major histocompatibility complex class I molecules on human lymphoid and myelomonocytic cells. J. Exp. Med. 186:1809.[Abstract/Free Full Text]
  22. Colonna, M., J. Samaridis, M. Cella, L. Angman, R. L. Allen, C. A. O’Callaghan, R. Dunbar, G. S. Ogg, V. Cerundolo, A. Rolink. 1998. Human myelomonocytic cells express an inhibitory receptor for classical and nonclassical MHC class I molecules. J. Immunol. 160:3096.[Abstract/Free Full Text]
  23. Allan, D. S., M. Colonna, L. L. Lanier, T. D. Churakova, J. S. Abrams, S. A. Ellis, A. J. McMichael, V. M. Braud. 1999. Tetrameric complexes of human histocompatibility leukocyte antigen (HLA)-G bind to peripheral blood myelomonocytic cells. J. Exp. Med. 189:1149.[Abstract/Free Full Text]
  24. Navarro, F., M. Llano, T. Bellon, M. Colonna, D. E. Geraghty, M. Lopez-Botet. 1999. The ILT2(LIR1) and CD94/NKG2A NK cell receptors respectively recognize HLA-G1 and HLA-E molecules co-expressed on target cells. Eur. J. Immunol. 29:277.[Medline]
  25. Uhrberg, M., N. M. Valiante, B. P. Shum, H. G. Shilling, K. Lienert-Weidenbach, B. Corliss, D. Tyan, L. L. Lanier, P. Parham. 1997. Human diversity in killer cell inhibitory receptor genes. Immunity 7:753.[Medline]
  26. Valiante, N. M., M. Uhrberg, H. G. Shilling, K. Lienert-Weidenbach, K. L. Arnett, A. D’Andrea, J. H. Phillips, L. L. Lanier, P. Parham. 1997. Functionally and structurally distinct NK cell receptor repertoires in the peripheral blood of two human donors. Immunity 7:739.[Medline]
  27. Shilling, H. G., K. Lienert-Weidenbach, N. M. Valiante, M. Uhrberg, P. Parham. 1998. Evidence for recombination as a mechanism for KIR diversification. Immunogenetics 48:413.[Medline]
  28. Gardiner, C. M., L. A. Guethlein, H. G. Shilling, M. Pando, W. H. Carr, R. Rajalingam, C. Vilches, P. Parham. 2001. Different NK cell surface phenotypes defined by the DX9 antibody are due to KIR3DL1 gene polymorphism. J. Immunol. 166:2992.[Abstract/Free Full Text]
  29. Rajalingam, R., C. M. Gardiner, F. Canavez, C. Vilches, P. Parham. 2001. Identification of seventeen novel KIR variants: fourteen of them from two non-Caucasian donors. Tissue Antigens 57:22.[Medline]
  30. Young, N. T., F. Canavez, M. Uhrberg, B. P. Shum, P. Parham. 2001. Conserved organization of the ILT/LIR gene family within the polymorphic human leukocyte receptor complex. Immunogenetics 53:270.[Medline]
  31. Khakoo, S. I., R. Rajalingam, B. P. Shum, K. Weidenbach, L. Flodin, D. G. Muir, F. Canavez, S. L. Cooper, N. M. Valiante, L. L. Lanier, et al 2000. Rapid evolution of NK cell receptor systems demonstrated by comparison of chimpanzees and humans. Immunity 12:687.[Medline]
  32. Lawlor, D. A., E. Warren, F. E. Ward, P. Parham. 1990. Comparison of class I MHC alleles in humans and apes. Immunol. Rev. 113:147.[Medline]
  33. Adams, E. J., S. Cooper, G. Thomson, P. Parham. 2000. Common chimpanzees have greater diversity than humans at two of the three highly polymorphic MHC class I genes. Immunogenetics 51:410.[Medline]
  34. Sambrook, J., E. F. Fritsch, T. Maniatis. 1989. Molecular Cloning: A Laboratory Manual Cold Spring Harbor Laboratory Press, Cold Spring Harbor.
  35. Devereux, J., P. Haeberli, O. Smithies. 1984. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 12:387.
  36. Saitou, N., M. Nei. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol. 4:406.[Abstract]
  37. Felsenstein, J.. 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39:783.
  38. Jakobsen, I. B., S. R. Wilson, S. Easteal. 1997. The partition matrix: exploring variable phylogenetic signals along nucleotide sequence alignments. Mol. Biol. Evol. 14:474.[Abstract]
  39. Nei, M., T. Gojobori. 1986. Simple methods for estimating the numbers of synonymous and nonsynonymous nucleotide substitutions. Mol. Biol. Evol. 3:418.[Abstract]
  40. Ota, T., M. Nei. 1994. Variance and covariances of the numbers of synonymous and nonsynonymous substitutions per site. Mol. Biol. Evol. 11:613.[Abstract]
  41. Jr Dennis, G., H. Kubagawa, M. D. Cooper. 2000. Paired Ig-like receptor homologs in birds and mammals share a common ancestor with mammalian Fc receptors. Proc. Natl. Acad. Sci. USA 97:13245.[Abstract/Free Full Text]
  42. Morin, P. A., J. J. Moore, R. Chakraborty, L. Jin, J. Goodall, D. S. Woodruff. 1994. Kin selection, social structure, gene flow, and the evolution of chimpanzees. Science 265:1193.[Abstract/Free Full Text]
  43. Chen, F. C., W. H. Li. 2001. Genomic divergences between humans and other hominoids and the effective population size of the common ancestor of humans and chimpanzees. Am. J. Hum. Genet. 68:444.[Medline]
  44. Kumar, S., S. B. Hedges. 1998. A molecular timescale for vertebrate evolution. Nature 392:917.
  45. Rajalingam, R., M. Hong, E. J. Adams, B. P. Shum, L. A. Guethlein, P. Parham. 2001. Short KIR haplotypes in Pygmy chimpanzee (Bonobo) resemble the conserved framework of diverse human KIR haplotypes. J. Exp. Med. 193:135.[Abstract/Free Full Text]
  46. Kimura, M.. 1980. A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J. Mol. Evol. 16:111.[Medline]



This article has been cited by other articles:


Home page
 Int ImmunolHome page
R. J. Lebbink, T. de Ruiter, G. J. A. Kaptijn, D. G. Bihan, C. A. Jansen, P. J. Lenting, and L. Meyaard
Mouse leukocyte-associated Ig-like receptor-1 (mLAIR-1) functions as an inhibitory collagen-binding receptor on immune cells
Int. Immunol., August 16, 2007; (2007) dxm071v1.
[Abstract] [Full Text] [PDF]

Home page
 Genome ResHome page
A. Varki and T. K. Altheide
Comparing the human and chimpanzee genomes: Searching for needles in a haystack
Genome Res., December 1, 2005; 15(12): 1746 - 1758.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
D. Belkin, M. Torkar, C. Chang, R. Barten, M. Tolaini, A. Haude, R. Allen, M. J. Wilson, D. Kioussis, and J. Trowsdale
Killer Cell Ig-Like Receptor and Leukocyte Ig-Like Receptor Transgenic Mice Exhibit Tissue- and Cell-Specific Transgene Expression
J. Immunol., September 15, 2003; 171(6): 3056 - 3063.
[Abstract] [Full Text] [PDF]

Home page
 BloodHome page
L. Borges, M. Kubin, and T. Kuhlman
LIR9, an immunoglobulin-superfamily-activating receptor, is expressed as a transmembrane and as a secreted molecule
Blood, February 15, 2003; 101(4): 1484 - 1486.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
T. Angata, S. C. Kerr, D. R. Greaves, N. M. Varki, P. R. Crocker, and A. Varki
Cloning and Characterization of Human Siglec-11. A RECENTLY EVOLVED SIGNALING MOLECULE THAT CAN INTERACT WITH SHP-1 AND SHP-2 AND IS EXPRESSED BY TISSUE MACROPHAGES, INCLUDING BRAIN MICROGLIA
J. Biol. Chem., June 28, 2002; 277(27): 24466 - 24474.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Canavez, F.
Right arrow Articles by Parham, P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Canavez, F.
Right arrow Articles by Parham, P.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS