A Human Homologue (BICD1) of theDrosophila Bicaudal-DGene

We previously isolated a cDNA fragment homologous to theDrosophila Bicaudal-Dgene (Bic-D) using a hybridization selection procedure with cosmids derived from the short arm of human chromosome 12. A PCR-mediated cDNA cloning strategy was applied to obtain the coding sequence of the human homologue (BICD1) and to generate a partial mouse (Bicdh1) cDNA. TheDrosophila Bicaudal-Dgene encodes a coiled coil protein, characterized by five α-helix domains and a leucine zipper motif, that forms part of the cytoskeleton and mediates the correct sorting of mRNAs for oocyte- and axis-determining factors during oogenesis. Analysis of the predicted amino acid sequence of theBICD1cDNA clones indicates that the sequence similarity is essentially limited to the amphipatic helices and the leucine zipper, but the conserved order of these domains suggests a similar function of the protein in mammalians. A database search further indicates the existence of a second human homologue on chromosome arm 9q and aCaenorhabditis eleganshomologue. Northern blot analysis indicates that both the human and the murine homologues produce an mRNA species of 9.5 kb expressed in brain, heart, and skeletal muscle and during mouse embryonic development. The conserved structural characteristics of theBICD1protein and its expression in muscle and especially brain suggest thatBICD1is a component of a cytoskeleton-based mRNA sorting mechanism conserved during evolution.     

Molecular cloning, characterization, and chromosomal localization of the mouse homologue of CD84, a member of the CD2 family of cell surface molecules

 CD84 is a member of the immunoglobulin gene superfamily (IgSF) with two Ig-like domains expressed primarily on B lymphocytes and macrophages. Here we describe the cloning of the mouse homologue of human CD84. Mouse CD84 cDNA clones were isolated from a macrophage library. The nucleotide sequence of mouse CD84 was shown to include an open reading frame encoding a putative 329 amino acid protein composed of a 21 amino acid leader peptide, two extracellular immunoglobulin (Ig)-like domains, a hydrophobic transmembrane region, and an 87 amino acid cytoplasmic domain. Mouse CD84 shares 57.3% amino acid sequence identity (88.7%, considering conservative amino acid substitutions) with the human homologue. Chromosome localization studies mapped the mouse CD84 gene to distal chromosome 1 adjacent to the gene for Ly-9, placing it close to the region where other members of the CD2 IgSF (CD48 and 2B4) have been mapped. Northern blot analysis revealed that the expression of mouse CD84 was predominantly restricted to hematopoietic tissues. Two species of mRNA of 3.6 kilobases (kb) and 1.5 kb were observed. The finding that the pattern of expression was restricted to the hematopoietic system and the conserved sequence of the mouse CD84 homologue suggests that the function of the CD84 glycoprotein may be similar in humans and mice. Received: 1 July 1998 / Revised: 31 August 1998     

The level of BMP4 signaling is critical for the regulation of distinct T-box gene expression domains and growth along the dorso-ventral axis of the optic cup

Background  

Polarised gene expression is thought to lead to the graded distribution of signaling molecules providing a patterning mechanism across the embryonic eye. Bone morphogenetic protein 4 (Bmp4) is expressed in the dorsal optic vesicle as it transforms into the optic cup. Bmp4 deletions in human and mouse result in failure of eye development, but little attempt has been made to investigate mammalian targets of BMP4 signaling. In chick, retroviral gene overexpression studies indicate that Bmp4 activates the dorsally expressed Tbx5 gene, which represses ventrally expressed cVax. It is not known whether the Tbx5 related genes, Tbx2 and Tbx3, are BMP4 targets in the mammalian retina and whether BMP4 acts at a distance from its site of expression. Although it is established that Drosophila Dpp (homologue of vertebrate Bmp4) acts as a morphogen, there is little evidence that BMP4 gradients are interpreted to create domains of BMP4 target gene expression in the mouse.     

Isolation of the Mouse Homologue of BRCA1 and Genetic Mapping to Mouse Chromosome 11

The BRCA1 gene is in large part responsible for hereditary human breast and ovarian cancer. Here we report the isolation of the murineBrca1homologue cDNA clones. In addition, we identified genomic P1 clones that contain most, if not all, of the mouseBrca1locus. DNA sequence analysis revealed that the mouse and human coding regions are 75% identical at the nucleotide level while the predicted amino acid identity is only 58%. A DNA sequence variant in theBrca1locus was identified and used to map this gene on a (Mus m. musculusCzech II × C57BL/KsJ)F1 × C57BL/KsJ intersubspecific backcross to distal mouse chromosome 11. The mapping of this gene to a region highly syntenic with human chromosome 17, coupled with Southern and Northern analyses, confirms that we isolated the murineBrca1homologue rather than a related RING finger gene. The isolation of the mouseBrca1homologue will facilitate the creation of mouse models for germline BRCA1 defects.     

Molecular genetics of the<Emphasis Type="Italic"> Alhambra</Emphasis> (<Emphasis Type="Italic"> Drosophila AF10</Emphasis>) complex locus of<Emphasis Type="Italic"> Drosophila</Emphasis>

The Alhambra ( Alh) gene is the Drosophila homologue of the human AF10 gene. AF10 has been identified as a fusion partner of MLL, a human homologue of the fly gene trithorax, in infant leukemias. The endogenous function of human AF10 is not known, but may be vital to its role in acute leukemia. This prompted us to analyse Alh function. We describe here the genetic organisation of the Alh locus in D. melanogaster. We show that an independent lethal complementation group encoding a muscle protein ( Mlp84B) is located within an Alh intron. We have already shown that the leucine zipper (LZ) domain of ALH activates several Polycomb group-responsive elements. We further demonstrate that the LZ domain on its own bears the Alh vital function, since it is necessary and sufficient for rescue of Alh mutant lethality. Finally, we demonstrate that, in contrast to a previous report, Alh does not affect position-effect variegation.Communicated by G. Reuter     

Sequence and Chromosomal Localization of Mouse Annexin XI

Mouse annexin XI (anx11)²was cloned from a macrophage cDNA library and characterized by genetic linkage mapping, DNA sequencing, and structural comparison with other annexins. TheAnx11gene localized to mouse chromosome 14 in close linkage with theRarb, Plau,andWnt5agenes near the centromere and 1.8 cM distal from theAnx7gene. The open reading frame was flanked by long, untranslated regions and encoded a 503-amino-acid protein with 93.1% identity to its human orthologue. Its 189-aa amino terminus corresponded to the widely expressed variant 1 of two possible, alternatively spliced forms. A previously described peptide fromAplysia brasilianawas identified as a closely related invertebrate homologue. Since annexin XI is known to be localized in the nucleus at certain stages of development, the identification of a region in tetrad repeats 3 and 4 resembling the “chromo box” domain may be relevant to a nuclear regulatory function of annexin XI. Knowledge of the mouse cDNA sequence and genetic map location will assist in the analysis of genomic organization and expression and provide a useful animal model to investigate gene function and hereditary phenotype for annexin XI.     

Molecular Cloning and Chromosomal Localization of the MouseGpr37Gene Encoding an Orphan G-Protein-Coupled Peptide Receptor Expressed in Brain and Testis

We report the cloning of the mouse ortholog of the humanGPR37gene, which encodes an orphan G-protein-coupled receptor highly expressed in brain tissues and homologous to neuropeptide-specific receptors ([20],Genomics 45:68–77;[45],Biochem. Biophys. Res. Commun. 233:559–567). The genomic organization of theGPR37gene is conserved in both mouse and human species with a single intron interrupting the receptor-coding sequence within the presumed third transmembrane domain. Comparative genetic mapping of theGPR37gene showed that it maps to a conserved chromosomal segment on proximal mouse chromosome 6 and human chromosome 7q31. The mouseGpr37gene contains an open reading frame coding for a 600-amino-acid protein 83% identical to the humanGPR37gene product. The predicted mouse GPR37 protein contains seven putative hydrophobic transmembrane domains, as well as a long (249 amino acid residues), arginine- and proline-rich amino-terminal extracellular domain, which is also a distinctive feature of the human GPR37 receptor. Northern blot analysis of mouse tissues withGpr37-specific probes revealed a main 3.8-kb mRNA and a much less abundant 8-kb mRNA, both expressed in the brain. A 3-kb mRNA is also expressed in the testis. Both the mouse and the humanGPR37genes may belong to a class of highly conserved mammalian genes encoding a novel type of G-protein-coupled receptor predominantly expressed in the brain.     

Location of the origin of replication for the 7.5-kbChlamydia trachomatis plasmid

The hypothetical origin of replication for the 7.5-kb plasmid common toChlamydia trachomatis is believed to be in a region of the plasmid that contains four 22-bp tandem repeats preceded by an A-T-rich region. To test this hypothesis, replication of plasmid DNA in metabolically active reticulate bodies of the Lymphogranuloma venereum biovar ofC. trachomatis was examined by electron microscopy. The results presented show that the origin of replication appears to be near the tandem repeats of pCHL2. In addition, replication of the 7.5-kb plasmid is unidirectional, and the copy number during replication is 7–10. The evidence presented suggests thatC. trachomatis has a homologue to theEscherichia coli dnaA gene and that this homologue might be involved in replication of theC. trachomatis 7.5-kb plasmid.     

Molecular Basis for the rhino (hr)Phenotype: A Nonsense Mutation in the MousehairlessGene

The hairless (hr) and rhino (hr^rh) mutations are autosomal recessive allelic mutations that map to mouse Chromosome 14. Both hairless and rhino mice have a number of skin and nail abnormalities and develop a striking form of total alopecia at approximately 3–4 weeks of age. The molecular basis of the hairless mouse phenotype was previously found to be the result of a murine leukemia proviral insertion in intron 6 of thehrgene that resulted in aberrant splicing. In this study, we report a 2-bp substitution in exon 4 of thehrgene in a second allele ofhr,rhino 8J (hr^rh-8J), leading to a nonsense mutation. These findings document the molecular basis of the rhino phenotype for the first time and suggest that rhino is a functional knock-out of thehrgene.     

Identification of replication origins in the genome of the methanogenic archaeon,<Emphasis Type="Italic"> Methanocaldococcus jannaschii</Emphasis>

Methanocaldococcus jannaschii has been notorious as an archaeon in which the replication origins are difficult to identify. Although extensive efforts have been exerted on this issue, the locations of replication origins still remain elusive 7 years after the publication of its complete genome sequence in 1996. Ambiguous results were obtained in identifying the replication origins of M. jannaschii based on all theoretical and experimental approaches. In the genome of M. jannaschii, we found that an ORF (MJ0774), annotated as a hypothetical protein, is a homologue of the Cdc6 protein. The position of the gene is at a global minimum of the x component of the Z curve, i.e., RY disparity curve, which has been used to identify replication origins in other Archaea. In addition, an intergenic region (694,540–695,226 bp) that is between the cdc6 gene and an adjacent ORF shows almost all the characteristics of known replication origins, i.e., it is highly rich in AT composition (80%) and contains multiple copies of repeat elements and AT stretches. Therefore, these lines of evidence strongly suggest that the identified region is a replication origin, which is designated as oriC1. The analysis of the y component of the Z curve, i.e., MK disparity curve, suggests the presence of another replication origin corresponding to one of the peaks in the MK disparity curve at around 1,388 kb of the genome.Communicated by G. Antranikian     

Identification and Characterization of a Seven Transmembrane Hormone Receptor Using Differential Display

Targeted mutagenesis analysis has shown that theCmybproto-oncogene, which encodes a sequence-specific DNA binding protein, is required for normal murine fetal liver erythropoiesis and myelopoiesis. To identify novel genes involved in hematopoiesis, differential display analysis was conducted using total liver RNA isolated from 14.5-day postcoitusCmybwildtype, heterozygous, and homozygous mutant littermates. Using 4 oligo(dT) 3′ primers and 5 arbitrary decamers as 5′ primers, 22 differentially expressed genes have been identified. Eight putatively novel genes were identified from 12 cDNAs that were sequenced. One gene, initially designated DD7A5-7, is primarily expressed in cells of the myeloid lineage. The full-length DD7A5-7 cDNA is 3239 nucleotides, encoding a putative protein of 931 amino acids. The protein is a member of a family of hormone receptors containing 7 transmembrane segments. The receptor also contains 7 epidermal growth factor-like (Egf-like) motifs at the amino terminal of the predicted protein. The gene is alternatively spliced, resulting in the deletion of one or more copies of theEgf-like motif. DD7A5-7 maps to mouse Chromosome 17 and is the putative homologue ofEMR1,a recently describedEgf-like module containing mucin-like hormone receptor with 7 transmembrane segments in humans. Our results indicate that theCmybmutant fetuses represent a unique resource for identifying genes involved in hematopoiesis.     

Genomic imprinting in ruminants: allele-specific gene expression in parthenogenetic sheep

Studies in the mouse have established that both parental genomes are essential for normal embryonic development. Parthenogenetic mouse embryos (which have two maternal genomes and no paternal genome), for example, are growth-retarded and die at early postimplantation stages. The distinct maternal and paternal contributions are mediated by genomic imprinting, an epigenetic mechanism by which the expression of certain genes is dependent on whether they are inherited from mother or father. Although comparative studies have established that many imprinted mouse (and rat) genes are allele-specifically expressed in humans as well (and vice versa), so far imprinting studies have not been performed in other mammalian species. When considering evolutionary theories of genomic imprinting, it would be important to know how widely it is conserved among placental mammals. We have investigated its conservation in a bovid ruminant, the domestic sheep, by comparing parthenogenetic and normal control embryos. Our study establishes that, like in the mouse, parthenogenetic development in sheep is associated with growth-retardation and does not proceed beyond early fetal stages. These developmental abnormalities are most likely caused by imprinted genes. We demonstrate that, indeed, like in mice and humans, the growth-related PEG1/MEST and Insulin-like Growth Factor 2 (IGF2) genes are expressed from the paternal chromosome in sheep. These observations suggest that genomic imprinting is conserved in a third, evolutionarily rather diverged group of placental mammals, the ruminants. Received: 13 May 1998 / Accepted: 16 July 1998     

Identification,expression and tissue distribution of a renalase homologue from mouse

FAD (flavin adenine dinucleotide)-dependent monoamine oxidases play very important roles in many biological processes. A novel monoamine oxidase, named renalase, has been identified in human kidney recently and is found to be markedly reduced in patients with end-stage renal disease (ESRD). Here, we reported the identification of a renalase homologue from mouse, termed mMAO-C (mouse monoamine oxidase-C) after the monoamine oxidase-A and -B (MAO-A and -B). This gene locates on the mouse chromosome 19C1 and its coding region spans 7 exons. The deuced amino acid sequences were predicted to contain a typical secretive signal peptide and a conserved amine oxidase domain. Phylogenetic analysis and multiple sequences alignment indicated that mMAO-C-like sequences exist in all examined species and share significant similarities. This gene has been submitted to the NCBI GenBank database (Accession number: DQ788834). With expression vectors generated from the cloned mMAO-C gene, exogenous protein was effectively expressed in both prokaryotic and eukaryotic cells. Recombinant mMAO-C protein was secreted out of human cell lines, indicating the biological function of its signal peptide. Moreover, tissue expression pattern analysis revealed that mMAO-C gene is predominantly expressed in the mouse kidney and testicle, which implies that kidney and testicle are the main sources of renalase secretion. Shortly, this study provides an insight into understanding the physiological and biological functions of mMAO-C and its homologues in endocrine.     

The expression pattern of a mouse doublesex-related gene is consistent with a role in gonadal differentiation

The signal for somatic sex determination in mammals, Caenorhabditis elegans and Drosophila melanogaster is chromosomal, but the overall mechanisms do not appear to be conserved between the phyla. However it has been found quite recently that the C. elegans sex-determining gene Mab-3 contains a domain highly homologous to the Drosophila sex-determining gene doublesex (dsx) and shares a similar role. These data suggest that at least some aspects of the regulation of sex determination might be conserved. In humans, a doublesex-related gene (DMRT1) was identified at less than 30 kb from the critical region for sex reversal on chromosome 9p24 (TD9). In order to get insights into the role of DMRT1 in sex determination/differentiation, we have isolated DMRT1 mouse homologue (Dmrt1) and analysed its expression pattern. The gene is expressed in the genital ridges of both sexes during the sex-determining switch and it shows male/female dimorphism at late stages of sex differentiation.     

Nucleotide Sequencing Analysis of the 146-Kilobase Segment around theIkBLandMICAGenes at the Centromeric End of the HLA Class I Region

To elucidate the complete gene structure and to identify new genes involved in the development ofHLAclass I antigen-associated diseases in the class I region of the human major histocompatibility complex on chromosome 6, a YAC clone (745D12) covering the 146-kb segment around theIkBLandMICAloci was isolated from a YAC library constructed from the B-cell line, BOLETH. A physical map of this region was constructed by isolation of overlapping cosmid clones derived from 745D12. Of these, five contiguous cosmids were chosen for DNA sequencing by the shotgun strategy to give a single contig of 146,601 bp from 2.8 kb telomeric of theIkBLgene to exon 6 ofMICA.This region was confirmed to contain five known genes,IkBL, BAT1, MICB, P5-1,andHLA-X(class I fragment), from centromere to telomere, and their exon–intron organizations were determined. The3.8-1homologue gene (3.8-1-hom) showing 99.7% identity with the3.8-1cDNA clone, which was originally isolated using the 3.8-kbEcoRI fragment between theHLA-54/Hand theHLA-Ggenes, was detected betweenMICAandMICBand was suggested to represent the cognate3.8-1genomic sequence from which the cDNA clone was derived. No evidence for the presence of expressed new genes could be obtained in this region by homology and EST searches or coding and exon prediction analyses. One TA microsatellite repeat spanning 2545 bases with as many as 913 repetitions was found on the centromeric side of theMICAgene and was indicated to be a potential hot spot for genetic recombination. The two segments of approximately 35 kb upstream of theMICAandMICBgenes showed high sequence homology (about 85%) to each other, suggesting that segmental genome duplication including theMICAandMICBgenes must have occurred during the evolution of the humanMHC.     

Chromosome mapping of the rod photoreceptor cGMP phosphodiesterase β-subunit gene in mouse and human: Tight linkage to the Huntington disease region (4p16.3)

The retinal degeneration mouse (gene symbol, rd) is an animal model for certain forms of human hereditary retinopathies. Recent findings of a nonsense mutation in the rd mouse PDE β-subunit gene (Pdeb) prompted us to investigate the chromosome locations of the mouse and human genes. We have utilized backcross analysis in mice to verify and define more precisely the location of the Pdeb locus 6.1 ± 2.3 cM distal of Mgsa on mouse chromosome 5. We have determined that the human gene (PDEB) maps to 4p16.3, very close to the Huntington disease (HD) region. Analysis of the comparative map for mice and humans shows that the mouse homologue of the HD gene will reside on chromosome 5. Linkage of the mouse Pdeb locus with other homologues in the human 4p16.3 region is maintained but gene order is not, suggesting at least three possible sites for the corresponding mouse HD gene.