The gene and the pseudogene for mouse p53 cellular tumor antigen are located on different chromosomes.

The chromosomal assignments of the two genes encoding the murine p53 cellular tumor antigen were determined by using a panel of mouse-Chinese hamster somatic cell hybrid clones and a mouse p53-specific cDNA clone. One gene, probably the functional member of the family, was found to be on chromosome 11. The other gene, which is probably a processed pseudogene, was assigned to chromosome 14. The potential relevance of these findings to documented cases of chromosome 11 trisomy are also discussed.     

Linkage map of two HLA-SB beta and two HLA-SB alpha-related genes: an intron in one of the SB beta genes contains a processed pseudogene

Three overlapping cosmid clones contain coding sequences for four HLA Class II genes, provisionally identified as two HLA-SB alpha and two HLA-SB beta genes. The genes are in the order beta, alpha, beta, alpha, inverted with respect to each other. One of the SB beta genes contains a 513 bp sequence that appears to be a processed pseudogene, flanked by direct 17 bp repeat sequences, in the intron upstream of the beta 1 exon. The pseudogene is homologous to a family of sequences of approximately 25-40 members, most of which are not on chromosome 6. A cDNA clone, highly homologous to the pseudogene, except for its 5' end, contains a normal poly(A) addition site and a poly(A) tail. The cDNA clone is homologous to a single-copy gene in both man and mouse, encoded on human chromosome 15. A search of published DNA sequences identified a mouse sequence, with about 77% similarity to the pseudogene sequence, in the negative strand of an intron in a mouse dihydrofolate reductase gene. The second SB beta gene does not contain the pseudogene sequence.     

Identification of two paralogous regions mapping to the short and long arms of human chromosome 2 comprising LIS1 pseudogenes

Reiner et al. (1995b) reported on the existence of a gene with a coding region virtually identical to LIS1, the gene responsible for Miller-Dieker lissencephaly. This gene, LIS2, was mapped to chromosome 2p11.2, and a related pseudogene, LIS2P, was mapped to 2q13-->q14. By sequencing genomic clones that were mapped by means of 2p and 2q-only hybrids, we now demonstrate the existence of two LIS1 processed pseudogenes mapping to 2p11.2 and 2q13 (PAFAH1P1 and PAFAH1P2, respectively). The two sequences appear to lie within larger paralogous regions and share a 98.6% degree of identity. Comparative mapping data by cytogenetic analysis on great apes indicate that the duplication of the genomic region comprising the LIS1 pseudogenes occurred in humans. We also demonstrate that the cDNA sequence shown as part of the LIS2 gene and marking its chromosome 2 specificity belongs to the 3' untranslated region of a different gene (C1orf6) that we mapped to 1q21 by FISH analysis.     

Identification and characterisation of a cytochrome P450 gene and processed pseudogene from an arachnid: the cattle tick, Boophilus microplus.

We isolated and sequenced the first known cytochrome P450 gene and pseudogene from an arachnid, the cattle tick, Boophilus microplus. Both the gene and pseudogene belong to the family CYP4, but a new subfamily, CYP4W, had to be created for these genes because they are substantially different to other CYP4 genes. The gene, CYP4W1, has greatest homology with CYP4C1 from a cockroach, Blaberus discoidalis. The predicted molecular weight of the protein encoded by CYP4W1 (63 KDa) is greater than that of the other CYP4 genes. The pseudogene, CYP4W1P, is probably a processed pseudogene derived from the functional gene CYP4W1. This is only the third CYP processed pseudogene to be identified. The pseudogene is 98% identical to the functional gene, CYP4W1, therefore we hypothesise that this pseudogene evolved recently from the functional gene. The CYP4 genes from arthropods have diverged from each other more than those of mammals; consequently the phylogeny of the arthropod genes could not be resolved.     

Physical localization and order of genes in the class I region of the bovine MHC

Fluorescence in situ hybridization (FISH) analyses were used to order 16 bacterial artificial chromosomes (BAC) clones containing loci from the bovine lymphocyte antigen (BoLA) class I and III regions of bovine chromosome 23 (BTA23). Fourteen of these BACs were assigned to chromosomal band locations of mitotic and pachytene chromosomes by single- and dual-colour FISH. Dual-colour FISH confirmed that class II DYA is proximal to and separated from BoLA class I genes by approximately three chromosome bands. The FISH results showed that tumour necrosis factor alpha (TNFA), heat shock protein 70 (HSP70.1) and 21 steroid dehydrogenase (CYP21) are closely linked in the region of BTA23 band 22 along with BoLA class I genes, and that male enhanced antigen (MEA) mapped between DYA and the CYP21/TNFA/HSP70.1 gene region. All BAC clones containing BoLA class I genes mapped distal to CYP21/TNFA/HSP70.1 and centromeric to prolactin (PRL). Myelin oligodendrocyte glycoprotein (MOG) was shown to be imbedded within the BoLA class I gene cluster. The cytogenetic data confirmed that the disrupted distribution of BoLA genes is most likely the result of a single large chromosomal inversion. Similar FISH results were obtained when BoLA DYA and class I BAC clones were mapped to discrete chromosomal locations on the BTA homologue in white-tailed deer, suggesting that this chromosomal inversion predates divergence of the advanced ruminant families from a common ancestor.     

Analysis of the genomic organisation of a small chromosome of Leishmania braziliensis M2903 reveals two genes encoding GTP-binding proteins, one of which belongs to a new G-protein family and is an antigen

Leishmania braziliensis M2903 contains a highly amplified small chromosome. This work is aimed at resolving its structural organization and determining whether this unusual chromosome contains specific genes encoding proteins with important functions in disease pathology or drug resistance. Our results show that the M2903 250-kb small chromosome contains LD1 sequences and has an inverted repeat structure. The LD1 sequences and two cDNAs (cDNA2 and cDNA53) were mapped on a cosmid contig, and the two cDNAs and the corresponding genomic fragments from the small chromosome were sequenced. The gene encoding cDNA2 predicts a putative GTP-binding protein with homology to other GTP-binding proteins only in the G-1 domain region; however, four other conserved motifs can be recognized. Sequence similarity to cDNA53 is located in at least five chromosomes, and its small chromosome copy is a pseudogene. An open reading frame downstream of the cDNA53 pseudogene predicts another GTP-binding protein that belongs to a new G-protein family with an unusual conserved GTP-binding domain and a newly characterized conserved sequence motif. A portion of this GTP-binding protein gene was studied previously in L. aethiopica as a recombinant antigen that reacts with human antibodies.     

小型猪CYP3A88基因克隆与其他动物的序列比较

目的克隆我国资源小型猪品系巴马香猪肝脏中的CYP3A88基因,并进行生物信息学分析。方法应用RACE（Rapid Amplification of cDNA Ends）技术对其全长进行扩增,测序,利用Internet和GenBank数据库对其序列进行生物信息学分析。结果首次克隆并鉴定了我国资源小型猪品系巴马香猪肝脏中CYP3A88（GenBank登录号：EF625347）的编码区,获得大小为1965bp的全长cDNA,编码区长为1512bp,编码503个氨基酸;比较核苷酸序列,与小型猪CYP3A39相似性高达94%,而与人等其它动物的CYP3A相似性则在86%以下;推导和分析氨基酸序列表明,与小型猪CYP3A其它成员（CYP3A39、CYP3A29、CYP3A22）进行对比,其相似性分别为92%,89%,80%,而将小型猪与人的CYP3A分别比对,小型猪CYP3A88与人CYP3A4相似性最高,为77%;对其二级结构预测,它可能含12个α螺旋,4个β折叠;经NCBI上的CDD程序分析可知,其39～491氨基酸区域为P4503A亚家族保守结构区域;经聚类分析,小型猪和狗的CYP3A与人有较近的进化关系;通过同源建模法对其在线建模,人CYP3A4晶体结构作为其模建模型,得到了其经典的三维结构。结论在猪CYP3A家族四个基因中,CYP3A88在序列和高级结构上均与人CYP3A4的最为相似。     

A novel human cytochrome P450 gene (P450IIB): chromosomal localization and evidence for alternative splicing.

We have isolated from a single human liver cDNA library two clones which are highly homologous (78% over the coding region) to the major phenobarbital-inducible P450 from rat (P450IIB1). This is the first direct demonstration of the presence of the P450IIB gene subfamily in humans. This subfamily is much less extensive than the rodent homologues, but does appear to contain at least two genes. Of the cDNA clones isolated one is apparently normally spliced, whereas the other lacks exon 8 and retains all or part of intron 5. Both clones contain transcribed Alu sequences. The human P450IIB gene has been located to chromosome 19q12----19q13.2 using a probe derived from intron 5, and is close to the CYP 2A locus encoding cytochrome P450IIA2. Restriction fragment length polymorphisms have been found with the enzymes BamHI and MspI which will enable linkage to be determined between these two loci.     

Comparison of genomic and cDNA sequences of guinea pig CYP2B18 and rat CYP2B2: absence of a phenobarbital-responsive enhancer module in the upstream region of the CYP2B18 gene

Potential mechanisms were investigated whereby CYP2B18, a cytochrome P450 gene exhibiting high constitutive expression but only low levels of phenobarbital-inducibility in the guinea pig liver, may be differentially regulated versus the highly inducible rat CYP2B2 gene. To comparatively assess potential regulatory sequences associated with CYP2B18, a guinea pig genomic library was screened enabling isolation of the CYP2B18 gene. The genomic screening process resulted in the identification of at least four closely-related CYP2B18 genes, designated here as CYP2B18A-D. Of these isolates, CYP2B18A exhibited sequence identical to that of the CYP2B18 cDNA. Further, the deduced amino acid sequence of the CYP2B18 cDNA was identical to that of N-terminal and internally-derived peptide sequences obtained in this investigation from CYP2B18 protein isolated from guinea pig liver. Genomic structural sequences were derived for CYP2B18A, together with the respective 5'-upstream and intronic regions of the gene. Comparison of the CYP2B18A and CYP2B2 gene sequences revealed the lack of repetitive LINE gene sequences in CYP2B18A, putative silencing elements that effect neighboring genes, although these sequences were present in both 5'-upstream and 3'-downstream regions of CYP2B2. We determined that the phenobarbital-responsive enhancer module was absent from the 5'-upstream region as well as the intronic regions of CYP2B18A gene. We hypothesize that the compromised phenobarbital inducibility of CYP2B18A stems from its lack of a functional phenobarbital responsive enhancer module.     

A pseudogene homologous to mouse transplantation antigens: Transplantation antigens are encoded by eight exons that correlate with protein domains

We have isolated about 30 to 40 different BALB/c mouse sperm DNA genomic clones that hybridize to cDNA clones encoding proteins homologous to transplantation antigens. One of these clones (27.1) was selected for sequence analysis because it was polymorphic in Southern blot analyses of the DNAs from BALB/c and CBA mice. A fragment of 5.7 kilobases of this clone was completely sequenced and found to contain a pseudogene whose sequence is highly homologous to the sequences of known transplantation antigens. Pseudogene 27.1 is split into eight exons that correlate with the structurally defined protein domains of transplantation antigens. Using Southern blot hybridization on the DNAs of different inbred mouse strains, we mapped the pseudogene to the Qa-2,3 region, a part of the Tla complex on chromosome 17 that is adjacent to the major histocompatibility complex. The Qa-2,3 region encodes lymphoid differentiation antigens homologous to the transplantation antigens in size, in peptide map profiles and in their association with β2-microglobulin. These mapping studies suggest that gene 27.1 may be a pseudogene for either a Qa antigen or an as yet undefined transplantation antigen. Accordingly, we may have isolated genes encoding lymphoid differentiation antigens of the Tla complex as well as those encoding transplantation antigens among the 30 to 40 different genomic clones isolated from our sperm library.     

Mapping and characterization of the mouse and human SS18 genes, two human SS18-like genes and a mouse Ss18 pseudogene.

We have previously isolated and characterized a mouse cDNA orthologous to the human synovial sarcoma associated SS18 (formerly named SSXT and SYT) cDNA. Here, we report the characterization of the genomic structure of the mouse Ss18 gene. Through in silico methods with sequence information contained in the public databases, we did the same for the human SS18 gene and two human SS18 homologous genes, SS18L1 and SS18L2. In addition, we identified a mouse Ss18 processed pseudogene and mapped it to chromosome 1, band A2-3. The mouse Ss18 gene, which is subject to extensive alternative splicing, is made up of 11 exons, spread out over approximately 45 kb of genomic sequence. The human SS18 gene is also composed of 11 exons with similar intron-exon boundaries, spreading out over about 70 kb of genomic sequence. One alternatively spliced exon, which is not included in the published SS18 cDNA, corresponds to a stretch of sequence which we previously identified in the mouse Ss18 cDNA. The human SS18L1 gene, which is also made up of 11 exons with similar intron-exon boundaries, was mapped to chromosome 20 band q13.3. The smaller SS18L2 gene, which is composed of three exons with similar boundaries as the first three exons of the other three genes, was mapped to chromosome 3 band p21. Through sequence and mutation analyses this gene could be excluded as a candidate gene for 3p21-associated renal cell cancer. In addition, we created a detailed BAC map around the human SS18 gene, placing it unequivocally between the CA-repeat marker AFMc014wf9 and the dihydrofolate reductase pseudogene DHFRP1. The next gene in this map, located distal to SS18, was found to be the TBP associated factor TAFII-105 (TAF2C2). Further analogies between the mouse Ss18 gene, the human SS18 gene and its two homologous genes were found in the putative promoter fragments. All four promoters resemble the promoters of housekeeping genes in that they are TATA-less and embedded in canonical CpG islands, thus explaining the high and widespread expression of the SS18 genes.     

Isolation and molecular characterization of bovine Rhesus-like transcripts and chromosome mapping of the relevant locus

The bovine erythrocyte membrane carries Rhesus (Rh)-like proteins. To obtain a bovine nucleotide probe, a cDNA library of foetal liver was constructed and screened with the human RhCE probe. Three clones (245 bp, 1012 bp and 1400 bp) were isolated and sequenced. They share a high degree of similarity (up to 73%) with Rh-like cDNAs of primates characterized so far and all of them were shown to contain a polymorphic microsatellite in their 3' untranslated region. Their sequences support the occurrence of different splicing isoforms transcribed from the same RH-like gene. One of the clones (1400 bp), which has a 134-nucleotide deletion causing a frameshift, is structurally similar to the human Rh4 cDNA isoform. Synteny mapping and genetic linkage analysis located the bovine RH-like locus on chromosome BTA2, on which none of the 10 previously mapped blood group systems are found. In situ hybridization mapped the RH-like locus to BTA2q45. No linkage was detected between the microsatellite and the only unmapped blood group system (locus F). These results strongly suggest that the putative bovine Rh-like polypeptides do not correspond to any previously described bovine blood group. Comparative studies of human and bovine maps clearly show that the human RH locus, which is located on HSA1p34-p36, and its bovine counterpart belong to a linkage group highly conserved between both species.     

Human chromosome 3: integration of 60 NotI clones into a physical and gene map

Sequence tagged sites generated for 60 NotI clones (NotI-STSs) from human chromosome 3-specific NotI-jumping and NotI-linking libraries were physically located using PCR screening of a radiation hybrid (RH) GeneBridge4 panel. The NotI map of chromosome 3 was generated using these RH-mapping data and those obtained earlier by FISH and sequencing of the corresponding NotI clones. The sequences of the NotI clones showed significant homologies with known genes and/or ESTs for 58 NotI-STSs (97%). These 58 NotI clones displayed 91-100% identity to 54 genes and 23 cDNA/EST clones. One known and two hypothetical protein-coding genes were localized for the first time and nine cDNA clones (unknown genes) were also carefully mapped only in this work. Three newly mapped genes are histone gene H1X (NR1-BK20C) and genes for hypothetical proteins THC1032178 and THC1024604 (NL1-243).     

Integrated genetic and physical map of the 1q31-->q32.1 region, encompassing the RP12 locus, the F13B and HF1 genes, and the EEF1AL11 and RPL30 pseudogenes.

The gene for autosomal recessive retinitis pigmentosa (RP12) with preserved para-arteriolar retinal pigment epithelium was previously mapped close to the F13B gene in region 1q31-->q32.1. A 4-Mb yeast artificial chromosome contig spanning this interval was constructed to facilitate cloning of the RP12 gene. The contig comprises 25 sequence-tagged sites, polymorphic markers, and single-copy probes, including five newly obtained probes. The contig orders the F13B and HF1 genes, as well as five expressed sequence tags, with respect to the integrated genetic map of this region. Homozygosity mapping resulted in refinement of the candidate gene locus for RP12 to a 1. 3-cM region. Currently, approximately 1 Mb of the contig is represented in P1-derived artificial chromosome (PAC) clones. Direct screening of a cDNA library derived from neural retina with PACs resulted in identification of the human elongation factor 1alpha pseudogene (EEF1AL11) and a human ribosomal protein L30 pseudogene (RPL30). A physical and genetic map covering the entire RP12 candidate gene region was constructed.     

Juxta-centromeric region of human chromosome 21 is enriched for pseudogenes and gene fragments

A physical map including four pseudogenes and 10 gene fragments and spanning 500 kb in the juxta-centromeric region of the long arm of human chromosome 21 is presented. cDNA fragments isolated from a selected cDNA library were characterized and mapped to the 831B6 YAC and to two BAC contigs that cover 250 kb of the region. An 85 kb genomic sequence located in the proximal region of the map was analyzed for putative exons. Four pseudogenes were found, including psiIGSF3, psiEIF3, psiGCT-rel whose functional copies map to chromosome 1p13, chromosome 2 and chromosome 22q11, respectively. The TTLL1 pseudogene corresponds to a new gene whose functional copy maps to chromosome 22q13. Ten gene fragments represent novel sequences that have related sequences on different human chromosomes and show 97-100% nucleotide identity to chromosome 21. These may correspond to pseudogenes on chromosome 21 and to functional genes in other chromosomes. The 85 kb genomic sequence was analyzed also for GC content, CpG islands, and repetitive sequence distribution. A GC-poor L isochore spanning 40 kb from satellite 1 was observed in the most centromeric region, next to a GC-rich H isochore that is a candidate region for the presence of functional genes. The pericentric duplication of a 7.8 kb region that is derived from the 22q13 chromosome band is described. We showed that the juxta-centromeric region of human chromosome 21 is enriched for retrotransposed pseudogenes and gene fragments transferred by interchromosome duplications, but we do not rule out the possibility that the region harbors functional genes also.     

A transcript map of an 800-kb region on human chromosome 11q13, part of the candidate region for SCA5 and BBS1

The exon-amplification method was used to identify putative transcribed sequences from an 800-kb region that includes the genes for phospholipase Cβ3 and PYGM on human chromosome 11q13. The clone contig consisted of ten cosmids, three bacterial artificial chromosomes, and one P1 artificial chromosome. A total of 83 exons were generated of which 23 were derived from known genes and expressed sequence tags (ESTs). Five different EST cDNA clones were identified and mapped on the contig. One is a homolog of the human p70S6 kinase (p70s6 k) gene whose function involves the translational regulation of ribosomal protein synthesis and thereby impacts on ribosomal biogenesis. The gene for p70s6 k is expressed universally, including within adipose cells and retina, and it could play a role in Bardet-Biedl syndrome type 1, which has been mapped to 11q13. Received: 22 July 1998 / Accepted: 24 August 1998