Coding Sequence, Genomic Organization, Chromosomal Localization, and Expression Pattern of the Signalosome ComponentCops2:The Mouse Homologue ofDrosophila Alien

TheDrosophila aliengene is highly homologous to the human thyroid receptor interacting protein, TRIP15/COPS2, which is a component of the recently identified signalosome protein complex. We identified the mouse homologue ofDrosophila alienthrough homology searches of the EST database. We found that the mouse cDNA encodes a predicted 443-amino-acid protein, which migrates at 50 kDa. The gene for the mousealienhomologue, namedCops2,includes 12 coding exons spanning 30 kb of genomic DNA on the central portion of mouse chromosome 2. MouseCops2is widely expressed in embryonic, fetal, and adult tissues beginning as early as E7.5. MouseCops2cDNA hybridizes to two mRNA bands in all tissues at 2.3 and 4 kb, with an additional 1.9-kb band in liver. Immunostaining of native and epitope tagged proteins localized the mouseCops2protein in both the cytoplasm and the nucleus, with larger amounts in the nucleus in some cells.     

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     

Human and mouse homologues of the Drosophila melanogaster tweety (tty) gene: a novel gene family encoding predicted transmembrane proteins

We have cloned cDNA for TTYH1, a human homologue of the Drosophila melanogaster tweety (tty) gene. The 450-residue predicted protein shows 27% amino acid sequence identity (51% similarity) to the Drosophila protein, which contains an additional C-terminal repetitive region. A second Drosophila homologue exhibits 42% identity (65% similarity) to the tty protein. Mouse (Ttyh1), macaque, and Caenorhabditis elegans homologues were also identified, and the complete coding sequence for the mouse gene was determined. The mouse protein is 91% identical to the human protein. Hydrophobicity analysis of the tty-related proteins indicates that they represent a new family of membrane proteins with five potential membrane-spanning regions. The yeast FTR1 and FTH1 iron transporter proteins and the mammalian neurotensin receptors 1 and 2 have a similar hydrophobicity profile, although there is no detectable sequence homology to the tty-related proteins. This suggests that the tweety-related proteins could be involved in transport of iron or other divalent cations or alternatively that they may be membrane-bound receptors. TTYH1 was mapped to chromosome 19q13.4 by FISH and by radiation hybrid mapping using the Stanford G3 panel.     

Identification of a novel fibroblast growth factor, FGF-23, preferentially expressed in the ventrolateral thalamic nucleus of the brain

We isolated mouse cDNA encoding a novel FGF (251 amino acids). As this is the 23rd documented FGF, we termed it FGF-23. FGF-23 has a hydrophobic amino terminus ( approximately 24 amino acids), which is a typical signal sequence. As expected, recombinant mouse FGF-23 was efficiently secreted by High Five insect cell-infected recombinant baculovirus containing the cDNA, indicating that FGF-23 is a secreted protein. We also isolated human cDNA encoding FGF-23 (251 amino acids), which is highly identical ( approximately 72% amino acid identity) to mouse FGF-23. Of human FGF family members, FGF-23 is most similar to FGF-21 and FGF-19 ( approximately 24% and approximately 22% amino acid identities, respectively). Human FGF-23 gene was localized on the chromosome 12p13 and found to be tandem linked (within 5.5 kb) to human FGF-6 gene. The expression of FGF-23 mRNA in mouse adult tissues was examined by real-time quantitative polymerase chain reaction. FGF-23 mRNA was mainly expressed in the brain and thymus at low levels. The localization of FGF-23 mRNA in the brain was examined by in situ hybridization. FGF-23 mRNA in the brain was found to be preferentially expressed in the ventrolateral thalamic nucleus. Therefore, FGF-23 is expected a unique FGF that plays roles in the function of the ventrolateral thalamic nucleus.     

Characterization of TH1 and CTSZ, two non-imprinted genes downstream of GNAS1 in chromosome 20q13

The clustering and coordinate regulation of many imprinted genes justifies positional searches for imprinted genes adjacent to known ones. We recently characterized a locus on 20q13, containing GNAS1, which has a highly complex imprinted expression pattern. In a search for neighbouring genes, we have now characterized a new gene, TH1, downstream of GNAS1. TH1 and GNAS1 are separated by more than 70 kb consisting largely of interspersed repetitive DNA. TH1 is the homologue of a gene that, in Drosophila, lies adjacent to the DNA repair gene mei-41. We have determined the full-length structures of human, mouse and Drosophila TH1. Though of unknown function, TH1 is highly conserved and widely expressed. Nonetheless, there is no similar Caenorhabditis elegans protein. We have also determined the complete genomic structures of human and Drosophila TH1. The Drosophila gene has five exons spanning 2.6 kb. The last three introns have precise equivalents in the human gene, which has 15 exons spanning 14 kb and is transcribed away from GNAS1. Using a single-nucleotide polymorphism in the 3' untranslated region, we have demonstrated biallelic TH1 expression in human fetal tissues, suggesting that, unlike GNAS1, TH1 is probably not imprinted. Immediately downstream of TH1 lies CTSZ, encoding the recently described cysteine protease, cathepsin Z. We have also elucidated the genomic structure of this gene; it has six exons spanning 12 kb and is oriented tail-to-tail with TH1, only 70 bp separating their polyadenylation sites. A polymorphism was again identified within the CTSZ 3' untranslated region and used to demonstrate biallelic expression in fetal tissues.     

The genomic organization,promoter position and expression profile of the mouse MRG15 gene

MORF4 (mortality factor on chromosome 4) and the novel related MRG (MORF4-related gene) gene family were identified when MORF4 was shown to induce senescence in a subset of tumor cell lines. The gene on chromosome 15 (MRG15) has high similarity to Drosophila MSL3, which is a component of the dosage compensation complex. MRG15 also has a chromodomain and may therefore function as a chromatin remodeling factor in a complex(es) involving a histone acetyltransferase, similar to MSL3. To complement our studies on human MRG15, we cloned and characterized the mouse MRG15 gene. Mouse MRG15 is expressed ubiquitously in adult tissues and at various embryonic stages, and expression in adult testis is higher than in other tissues. MRG15-b, which is an alternatively spliced form of MRG15-a and has a 39-amino-acid insertion in the chromodomain, is also expressed in all mouse tissues examined and localizes to the nucleus of cells. It is possible that MRG15-b may lack the function of the chromodomain because of the additional amino acids and could potentially be the equivalent of the human MORF4 in the mouse. The mouse MRG15 gene is composed of twelve exons and spans over 24 kb DNA. Using luciferase constructs we have determined that there is a functional promoter sequence 1.8 kb upstream of the ATG start codon. This region contains no TATA box but has GC-rich regions, consistent with the ubiquitous expression we have observed.     

TTYH2, a human homologue of the Drosophila melanogaster gene tweety, is located on 17q24 and upregulated in renal cell carcinoma

Using differential display PCR, we identified a novel gene upregulated in renal cell carcinoma. Characterization of the full-length cDNA and gene revealed that the encoded protein is a human homologue of the Drosophila melanogaster Tweety protein, and so we have termed the novel protein TTYH2. The orthologous mouse cDNA was also identified and the predicted mouse protein is 81% identical to the human protein. The encoded human TTYH2 protein is 534 amino acids and, like the other members of the tweety-related protein family, is a putative cell surface protein with five transmembrane regions. TTYH2 is located at 17q24; it is expressed most highly in brain and testis and at lower levels in heart, ovary, spleen, and peripheral blood leukocytes. Expression of this gene is upregulated in 13 of 16 (81%) renal cell carcinoma samples examined. In addition to a putative role in brain and testis, the over-expression of TTYH2 in renal cell carcinoma suggests that it may have an important role in kidney tumorigenesis.     

cDNA cloning and localization of the mouse leukosialin gene (Ly48) to chromosome 7

Mouse leukosialin, previously known as the 3E8 antigen, is expressed primarily on cells of the hematopoietic and lymphoid lineages and is shown to be the mouse homologue to the human leukosialin/sialophorin and rat W3/13 molecules. A partial leukosialin cDNA clone was isolated via cross-species hybridization with a portion of a human leukosialin cDNA. This mouse cDNA clone was used to demonstrate that the leukosialin isoforms are encoded by a single mRNA species of approximately 4.2 kilobases (kb) and that the leukosialin gene is located on chromosome 7. Based on these results, mouse leukosialin is given the designation Ly48.The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession number M30693.     

Drosophila RNA polymerase II elongation factor DmS-II has homology to mouse S-II and sequence similarity to yeast PPR2.

DmSII is a Drosophila RNA polymerase II elongation factor which suppresses pausing by RNA polymerase II at specific sites on double stranded templates. Using antibodies produced against the purified protein, a Drosophila cDNA expression library was screened and a cDNA was isolated which encoded a portion of DmSII. When this cDNA was used to probe Kc cell mRNA the predominant species was found to be 1.4 kb in length. The original cDNA was used to screen a Drosophila Kc cell cDNA library resulting in the isolation of a 1.4 kb cDNA which was then sequenced. The deduced protein sequence for DmSII exhibited high similarity to mouse SII protein sequence. In addition, significant sequence similarity was found with the protein encoded by the yeast gene PPR2, which is involved in regulation of URA4 gene expression. The comparison of amino acid sequences suggests that DmSII is comprised of two domains homologous to mouse SII separated by a flexible, serine rich region of low homology. The shorter yeast protein has sequence similarity only to the carboxy terminal domain.