Evolution of genes for allelic and isotypic forms of immunoglobulin kappa chains and of the genes for T-cell receptor beta chains in rabbits

New insights into the evolution of the families of genes encoding immunoglobulins and T-cell receptors of rabbits (Oryctolagus cuniculus) have come from molecular genetic studies. In contrast to human and mouse, rabbits were shown to have two genes for the constant region of immunoglobulin light chains (C kappa 1 and C kappa 2 isotypes) and complex allelic variants of K1 (allotypes). Although K1 allotype protein sequences differed at up to 41% of the amino acid positions, 3' untranslated, 5', and 3' flanking regions were conserved, and in the coding regions 78-80% of the codons with differences had replacement changes. Proportions of silent changes and changes in noncoding regions were comparable. Thus, in spite of their markedly different protein sequences, the K1b4, b5, and b9 allotypes appeared to be products of allelic genes. Molecular genetic analyses suggested that they may have undergone rapid divergence after an ancestral K2-like gene duplicated. Some rabbits were found to have two similar T-cell receptor C beta genes as do humans and many strains of mice, but others appeared to have three different C beta. In addition, we found allotypic forms of C beta. Some of the C beta allotypic differences occurred at positions where analogous C kappa allotypic differences were found. We also found V beta in mouse and human that were more similar to rabbit V beta than closely linked rabbit genes were to each other. This contrasts with rabbit immunoglobulin VH gene sequences that reflect concerted evolution. The data suggested that T-cell receptor V beta genes duplicated prior to mammalian radiation.     

Comprehensive analysis of keratin gene clusters in humans and rodents

Here, we present the comparative analysis of the two keratin (K) gene clusters in the genomes of man, mouse and rat. Overall, there is a remarkable but not perfect synteny among the clusters of the three mammalian species. The human type I keratin gene cluster consists of 27 genes and 4 pseudogenes, all in the same orientation. It is interrupted by a domain of multiple genes encoding keratin-associated proteins (KAPs). Cytokeratin, hair and inner root sheath keratin genes are grouped together in small subclusters, indicating that evolution occurred by duplication events. At the end of the rodent type I gene cluster, a novel gene related to K14 and K17 was identified, which is converted to a pseudogene in humans. The human type II cluster consists of 27 genes and 5 pseudogenes, most of which are arranged in the same orientation. Of the 26 type II murine keratin genes now known, the expression of two new genes was identified by RT-PCR. Kb20, the first gene in the cluster, was detected in lung tissue. Kb39, a new ortholog of K1, is expressed in certain stratified epithelia. It represents a candidate gene for those hyperkeratotic skin syndromes in which no K1 mutations were identified so far. Most remarkably, the human K3 gene which causes Meesmann's corneal dystrophy when mutated, lacks a counterpart in the mouse genome. While the human genome has 138 pseudogenes related to K8 and K18, the mouse and rat genomes contain only 4 and 6 such pseudogenes. Our results also provide the basis for a unified keratin nomenclature and for future functional studies.     

Gene sequencing and tissue expression of unknown isoforms of an angiotensin II type 2 receptor interacting protein, ATIP, in the rat

To investigate the expression of the unknown angiotensin II type 2 receptor interacting protein (ATIP) isoforms in the rat we used the known sequences of human and mouse ATIP to design sequencing primers to enable us to sequence rat ATIP3 and ATIP4. Exon 4, which is present in human but not mouse ATIP, was not identified in the coding region of rat ATIP. The expression levels of these genes in a range of rat tissues were examined, and we concluded that there is little similarity in the relative tissue distribution of the various ATIP isoforms in rat and human.     

Comparative human/murine sequence analysis of the common eliminated region 1 from human 3p21.3

Interspecies sequence comparison offers an effective approach to identify conserved elements that might have functional importance. We compared 1.32 Mb of C3CER1 (referred also as CER1) from human Chromosome (Chr) 3p21.3 to its orthologous regions on mouse Chr 9F. The corresponding mouse region was found divided into two blocks, but their gene content and gene positions were highly conserved between human and mouse. We observed that two orthologous mouse genes (Xtrp3s1 and Cmkbr1) were duplicated, and this resulted in two additional expressed mouse genes (Xtrp3 and Cmkbr111). We also recognized a large number of conserved elements that were neither exons, CpG islands, nor repeats. We further identified and characterized five novel orthologous mouse genes (Kiaa0028, Xtrp3s1, Fyco1, Tmem7, and Lrrc2).     

Structural analysis of 5'-flanking regions of rat, mouse and human renin genes reveals the presence of a transposable-like element in the two mouse genes

The two renin genes of the mouse (Ren1 and Ren2) are expressed at different levels in the submaxillary gland (SMG). In contrast to mice, there is no detectable renin gene expression in the rat SMG. To determine the molecular basis for these different levels of renin expression, we have compared the 5'-flanking regions of the rat and mouse genes. The sequence of mouse, but not rat, genes reveals the presence in Ren1 and Ren2 of a large insertion, probably a new class of transposable elements. A second, apparently unrelated shorter insertion is present only in Ren2. Otherwise, the mouse and rat 5'-flanking sequences are well conserved and resemble the corresponding region of the human Ren gene, indicating that the insertions occurred after the separation of the rat and mouse species but before the duplication of the mouse Ren gene. We suggest that these structural differences may have a role in the differential expression of the Ren genes in mice and other animals.     

Phylogenetic tests of the hypothesis of block duplication of homologous genes on human chromosomes 6, 9, and 1

There are 10 gene families that have members on both human chromosome 6 (6p21.3, the location of the human major histocompatibility complex [MHC]) and human chromosome 9 (mostly 9q33-34). Six of these families also have members on mouse chromosome 17 (the mouse MHC chromosome) and mouse chromosome 2. In addition, four of these families have members on human chromosome 1 (1q21-25 and 1p13), and two of these have members on mouse chromosome 1. One hypothesis to explain these patterns is that members of the 10 gene families of human chromosomes 6 and 9 were duplicated simultaneously as a result of polyploidization or duplication of a chromosome segment ("block duplication"). A subsequent block duplication has been proposed to account for the presence of representatives of four of these families on human chromosome 1. Phylogenetic analyses of the 9 gene families for which data were available decisively rejected the hypothesis of block duplication as an overall explanation of these patterns. Three to five of the genes on human chromosomes 6 and 9 probably duplicated simultaneously early in vertebrate history, prior to the divergence of jawed and jawless vertebrates, and shortly after that, all four of the genes on chromosomes 1 and 9 probably duplicated as a block. However, the other genes duplicated at different times scattered over at least 1.6 billion years. Since the occurrence of these clusters of related genes cannot be explained by block duplication, one alternative explanation is that they cluster together because of shared functional characteristics relating to expression patterns.      

The immunoglobulin lambda locus in rat consists of two C lambda genes and a single V lambda gene

The immunoglobin lambda locus of the rat has been studied. Germ-line V lambda and C lambda genes were isolated from recombinant-phage libraries and characterized by nucleotide sequencing. The results showed that the lambda locus of the rat contains one single V lambda gene and two C lambda genes, thus representing one of the least complex lambda loci so far characterized. The two C lambda genes are separated by a spacer approx. 3 kb long. Two J segments are located at the 5' side of each C lambda gene. One of the C lambda genes (C lambda 1) probably represents a pseudogene, as the J lambda 1 segments have non-functional recombination and splice signals. The organization of the rat lambda locus resembles that of mouse, except that only one cluster is present in the rat. Thus since the evolutionary separation of the rat and mouse species ten MYR ( = 10(6) years) ago, either one cluster has been lost from the rat, or duplicated in the mouse.     

Comparative FISH mapping of Gab1 and Gab2 genes in human, mouse and rat.

Gab1 and Gab2 are members of the Gab family which act as adapters for transmitting various signals in response to stimuli through cytokine and growth factor receptors, and T- and B-cell antigen receptors. We determined chromosome locations of the two genes in human, mouse and rat by fluorescence in situ hybridization. The Gab1 gene was localized to chromosome 4q31.1 in human, 8C3 in mouse and 19q11.1--> q11.2 in rat, and the Gab2 gene was located on chromosome 11q13.4-->q13.5 in human, 7E2 in mouse and 1q33.2-->q33.3 in rat. All human, mouse and rat Gab1 and Gab2 genes were localized to chromosome regions where conserved homology has been identified among the three species.     

Tollip, a negative regulator of TLR-signalling, is encoded by twin genes in salmonid fish

The factor Tollip is known to dampen TLR2- and TLR4-mediated signalling in mammals. No negative regulator of the piscine TLR-signalling cascade has been described so far, albeit a sizable collection of factors contributing to this ancient pathogen-sensing system are known from fish to date. We identified two closely related Tollip-encoding genes in Atlantic salmon (Salmo salar) and the respective ortholog mRNA molecules in rainbow trout (Oncorhynchus mykiss). The salmonid Tollip genes are segmented into 6 exons, similar to the human orthologous gene. The protein-encoding sequences are homologous to >97% among the twin factors and also between the species. Both encoded proteins contain a C2 domain and an ubiquitin system component, which are also characteristic features of the mammalian Tollip factor. We analysed the expression of these genes in trout. Both Tollip-encoding genes are ubiquitously and also equally expressed, as indicated by similar mRNA concentrations of both factors in any one tissue. Tollip expression was found to be up-regulated by viral infection. Our data suggest that the Tollip genes were duplicated before salmon and trout were evolutionary separated. Moreover, pathways dampening the activity of the TLR-cascade may have been conserved from lower vertebrates to mammals since Tollip, as a respective key factor has been highly conserved from fish to human.     

Nonspecific esterases of Mus musculus

Seventeen genes controlling the expression of carboxylic ester hydrolases, commonly known as esterases, have been identified in the mouse Mus musculus. Seven esterase loci are found on chromosome 8, where two clusters of esterase loci occur. It seems probable that the genes within these clusters have arisen from a common ancestral gene by tandem duplication. Close linkage of esterase genes is also found in the rat, rabbit, and prairie vole. Some mouse esterases appear to be homologous with certain human esterases. The function of these nonspecific enzymes is still unknown.     

Characterization of the Bex gene family in humans, mice, and rats

To better understand the development of ventral mesencephalic dopamine neurons, we performed subtractive hybridization screens to find ventral mesencephalic genes expressed at rat embryonic day 10 when these neurons begin to differentiate. The most commonly identified genes in these screens were members of the Bex (Brain expressed X-linked) gene family, rat Bex1 (Rex3), and a novel gene, rat Bex4. After identifying these genes, we then sought to characterize the Bex gene family. Two additional novel Bex genes (human Bex5 and mouse Bex6) were discovered through genomic databases. Bex5 is present in humans and monkeys, but not rodents, while Bex6 exists in mice, but not humans. Bex4 and Bex5 are localized to the X chromosome, are expressed in brain, and are similar in sequence. Bex4 and Bex5 are 54% and 56% identical to human Bex3 (pHGR74, NADE). Mouse Bex6 is on chromosome 16 and is 67% identical to mouse Bex4. Human Bex gene expression was studied with tissue expression arrays probed with specific oligonucleotides. Human Bex1 and Bex2 have similar expression patterns in the central nervous system with high levels in pituitary, cerebellum, and temporal lobe, and Bex1 is widely expressed outside of the central nervous system with high expression in the liver. Human Bex4 is highly expressed in heart, skeletal muscle, and liver, while Bex3 and Bex5 are more widely expressed. The subcellular localization of the Bex proteins varies from nuclear (rat Bex1) to cytoplasmic (rat Bex3, human Bex5, and mouse Bex6) and to both nuclear and cytoplasmic (rat Bex2 and rat Bex4). Rat Bex3, rat Bex4, human Bex5, and mouse Bex6 are degraded by the proteasome, while rat Bex1 or Bex2 are not. Rat Bex3 protein can likely bind transition metals through a histidine-rich domain. Because this gene family was originally named Bex and because these genes are unified by sequence similarity and gene structure, we believe the Bex nomenclature should prevail over nomenclature based on function (NADE) that has not been extended to the other Bex genes. We conclude that the Bex gene family members are highly homologous but differ in their expression patterns, subcellular localization, and degradation by the proteasome.     

A genome-wide survey on basic helix-loop-helix transcription factors in rat and mouse

The basic helix-loop-helix (bHLH) proteins play essential roles in a wide range of developmental processes in higher organisms. bHLH family members have been identified in over 20 organisms, including nematode, fruit fly, and human. Our study identified 114 rat and 14 additional mouse bHLH members in rat and mouse genomes, respectively. Phylogenetic analyses revealed that both rat and mouse had 49, 26, 15, 4, 12, and 4 bHLH members in groups A, B, C, D, E, and F, respectively. Only the rat Mxi1 gene has two copies in the genome. All other rat bHLH genes and all mouse bHLH genes are single-copy genes. The chromosomal distribution pattern of mouse, rat, and human bHLH genes suggests the emergence of some bHLH genes through gene duplication, which probably happened at least before the divergence of vertebrates from invertebrates. The present study provides useful information for future studies using rat as a model animal for mammalian development. X. Zheng and Y. Wang are jointly first authors. An erratum to this article can be found at