Assignment of human asialoglycoprotein receptor gene (ASGR1) to chromosome 17p11-13.
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Animals↗
Biomedical subjects
R L Eddy
Publications and source records attributed to R L Eddy.
Assignment of the human prohibitin gene (PHB) to chromosome 17 and identification of a DNA polymorphism.
Prohibition is a recently identified antiproliferative protein whose exact role in the cell is under investigation. To determine the human chromosomal location of the prohibition gene (PHB) and whether this site corresponds to that of any suspected tumor suppressor gene, we have analyzed DNA from three sources by hybridization analysis: mouse--human hybrid cell lines, hybrid cell lines containing portions of human chromosomes, and human metaphase chromosomes in situ. All three techniques confirm a location in the region 17q21-q22, a region genetically linked to early-onset human breast cancer. Further analysis will be required to establish the significance of this relationship; Southern hybridizations show a polymorphic EcoRI site that may be useful for this purpose.
Blotting, Southern↗
Cloning of human heparan sulfate proteoglycan core protein, assignment of the gene (HSPG2) to 1p36.1----p35 and identification of a BamHI restriction fragment length polymorphism.
We have isolated a cDNA coding for the core protein of the large basement membrane heparan sulfate proteoglycan (HSPG) from a human fibrosarcoma cell (HT1080) library. The library was screened with a mouse cDNA probe and one clone obtained, with a 1.5-kb insert, was isolated and sequenced. The sequence contained an open reading frame coding for 507 amino acid residues with a 84% identity to the corresponding mouse sequence. This amino acid sequence contained several cysteine-rich internal repeats similar to those found in component chains of laminin. The HSPG cDNA clone was used to assign the gene (HSPG2) to the p36.1----p35 region of chromosome 1 using both somatic cell hybrid and in situ hybridization. In the study of the polymorphisms of the locus, a BamHI restriction fragment length polymorphism was identified in the gene. This polymorphism displayed bands of 23 and 12 kb with allele frequencies of 76 and 24%, respectively.
Amino Acid Sequence↗
Assignment of the human small inducible cytokine A2 gene, SCYA2 (encoding JE or MCP-1), to 17q11.2-12: evolutionary relatedness of cytokines clustered at the same locus.
The JE gene, cloned from platelet-derived growth factor (PDGF)-treated mouse 3T3 cells, was the first PDGF-inducible gene to be described. Its human homolog (gene name "small inducible cytokine A2" [SCYA2]) encodes the monocyte specific chemotactic factor MCP-1 (or MCAF) which is structurally related to a recently described family of cytokines. By a combination of in situ hybridization and study of somatic cell hybrids, we have assigned the human SCYA2 gene to 17q11.2-12, the locus to which other members of this family have been mapped. We have also reconstructed a phylogenetic tree relating the members of this family to each other and to their murine homologs which suggests that these genes arose by duplication and divergence prior to the murine/human divergence. Four of the five members of this subfamily have now been assigned to the same locus (and the fifth to chromosome 17), while several of the members of a related gene family have been assigned to 4q. We propose that the two subfamilies be designated the 17q and 4q subfamilies.
Amino Acid Sequence↗
Regional assignment of two genes of the human branched-chain alpha-keto acid dehydrogenase complex: the E1 beta gene (BCKDHB) to chromosome 6p21-22 and the E2 gene (DBT) to chromosome 1p31.
Maple syrup urine disease (MSUD) is caused by the deficiency of the mitochondrial branched-chain alpha-keto acid dehydrogenase complex. The multienzyme complex is a macromolecule (Mr 4 X 10(6] consisting of at least six distinct subunits. In this study, the human E1 beta gene (BCKDHB) has been localized to human chromosome 6 by hybrid somatic cell analysis, and regionally assigned to chromosome bands 6p21-22 by in situ hybridization. The E2 gene (DBT), which was previously localized to chromosome 1, is regionally assigned to the chromosome band 1p31 also by in situ hybridization. Localization of the E1 beta gene to chromosome 6p21-22 assigns another major human disease locus to a region that contains several important genes, including the major histocompatability complex, tumor necrosis factor, and heat-shock protein HSP70. Mapping of the E1 beta and the E2 genes may provide information for the linkage analysis of MSUD families with mutations in these two loci.
3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
cDNA cloning and chromosomal assignment of the endothelin 2 gene: vasoactive intestinal contractor peptide is rat endothelin 2.
Four members of the endothelin family of vasoactive and mitogenic peptides have been identified: human endothelins 1, 2, and 3 (ET1, ET2, and ET3, respectively) and mouse vasoactive intestinal contractor (VIC). To characterize the mRNA encoding ET2, a 192-bp fragment of the ET2 gene, amplified by the polymerase chain reaction from human genomic DNA, was used to screen cell lines and tissues for ET2 gene expression. ET2 mRNA was detected in a cell line (HTB119) derived from a human lung small cell carcinoma, and an ET2 cDNA was cloned from a cDNA library prepared from HTB119 mRNA. DNA prepared from human-mouse somatic hybrid cell lines was used to assign the gene encoding ET2 (EDN2) to the 1p21----1pter region of chromosome 1, demonstrating that EDN2 is not linked to genes encoding ET1 (EDN1; chromosome 6) and ET3 (EDN3; chromosome 20). Southern blot hybridization revealed a single gene in human and rat genomes that hybridized with the ET2 gene fragment, and the rat gene was cloned. The endothelin peptide encoded by the rat gene differed from ET2 at 1 of 21 residues and was identical to mouse VIC. We conclude that VIC is the mouse and rat analogue of the human ET2 gene.
Amino Acid Sequence↗
The human connexin gene family of gap junction proteins: distinct chromosomal locations but similar structures.
Connexins are protein subunits that constitute gap junction channels. Two members of this gene family, connexin43 (Cx43) and connexin32 (Cx32), are abundantly expressed in the heart and liver, respectively. Human genomic DNA analysis revealed the presence of two loci for Cx43: an expressed gene and a processed pseudogene. The expressed gene (GJA1) was mapped to human chromosome 6 and the pseudogene (GJA1P) to chromosome 5. To determine whether Cx32 was linked to Cx43, somatic cell hybrids were analyzed by polymerase chain reaction and hybridization, resulting in the assignment of the gene for Cx32 (GJB1) to the X chromosome at Xp11----q22. Comparison of the structures of connexin genes suggests that members of this multigene family arose from a single precursor, but evolved to distinct chromosomal locations.
Base Sequence↗
Localization of the dihydrolipoamide branched-chain transacylase gene (DBT) of the human branched-chain keto acid dehydrogenase complex to chromosome 1.
The gene coding for the transacylase subunit (DBT) of the human branched-chain keto acid dehydrogenase complex was localized to chromosome 1 by probing panels of human x mouse chromosome hybrids with an E2 cDNA amplified by the polymerase chain reaction. Additional data with two hybrids containing chromosome 1 fragments suggest that the DBT gene is located on the short arm (1pter----p21) of the chromosome.
3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
Epidermal type I transglutaminase (TGM1) is assigned to human chromosome 14.
Human epidermal type I transglutaminase coexists in keratinocytes with another cross-linking enzyme, tissue type II transglutaminase. There are at least five different forms of the enzyme in mammals. Gene mapping studies allowed us to determine whether the different transglutaminases are products of the same gene or separate genes. The gene encoding factor XIII subunit a transglutaminase (F13A1) was previously assigned to human chromosome 6, p24----p25. We demonstrate using somatic cell hybrids that the human epidermal type I transglutaminase gene (gene symbol is designated TGM1) is located on human chromosome 14, providing evidence that at least two human transglutaminases are encoded by separate genes.
Animals↗
Three epidermal and one simple epithelial type II keratin genes map to human chromosome 12.
We have localized the genes which encode the human type II epidermal keratins K5, K6a, and K6b and the simple epithelial keratin K7 (KRT5, KRT6A, KRT6B, and KRT7, respectively) to chromosome 12 using Southern blot analysis of somatic cell hybrids. In addition, we have sublocalized the genes for K6a and K7 to bands 12q12----q14 on the long arm of this chromosome by in situ hybridization of metaphase chromosomes.
Blotting, Southern↗
Identification of a new endothelial cell growth factor receptor tyrosine kinase.
A new growth factor receptor tyrosine kinase (RTK) gene (designated KDR) has been cloned from a human endothelial cell cDNA library. The gene was identified using the polymerase chain reaction (PCR) and degenerate oligonucleotide primers complementary to conserved tyrosine kinase domains that flank the insert domain, characteristic of known type III RTKs [e.g. platelet-derived growth factor receptor (PDGF-R), colony-stimulating-1 receptor (CSF-1-R), fibroblast growth factor receptor (FGF-R) and ckit]. The DNA product from PCR was then used as a probe to isolate larger DNA segments encoding the receptor from the cDNA library. The predicted amino acid sequence contained multiple characteristics (i.e. an ATP-binding site, a membrane-spanning region, split tyrosine kinase regions) typical of a type III receptor tyrosine kinase. The KDR gene is expressed as a 7.0 kb transcript, and is localized to human chromosome 4.
Amino Acid Sequence↗
A human gene homologous to the formin gene residing at the murine limb deformity locus: chromosomal location and RFLPs.
The murine limb deformity (ld) locus resides on mouse chromosome 2 and gives rise to a recessively inherited, characteristic limb deformity/renal aplasia phenotype. In this locus in the mouse, a gene, termed the "formin" gene, has been identified which encodes an array of differentially processed transcripts in both adult and embryonic tissues. A set of these transcripts are disrupted in independent mutant mouse ld alleles. We wish to report the isolation of a human genomic clone which is homologous to the mouse formin gene by virtue of sequence comparison and expression of conserved exons. Among human fetal tissues analyzed, the kidney appears to be a major site of expression. This human gene, LD, maps to chromosome 15q11----qter in mouse human somatic cell hybrids and, specifically, to 15q13----q14 by chromosomal in situ hybridization. This localization establishes both LD and beta 2-microglobulin as syntenic genes on mouse chromosome 2 and human chromosome 15 and implies the interspecies conservation of the region between them. In addition, we identify in the human locus two frequently occurring DNA polymorphisms which can be used to test the linkage of LD to known human dysmorphoses.
Alleles↗
The human neurokinin A (substance K) receptor. Molecular cloning of the gene, chromosome localization, and isolation of cDNA from tracheal and gastric tissues.
Neurokinin A (substance K) is a peptide neurotransmitter of the tachykinin family with potential as a major mediator in human airway and gastrointestinal tissues. Neurokinin A acts via a receptor (the NK-2 receptor) believed to be localized on smooth muscle cells and pharmacologically coupled to a GTP-binding protein. To characterize the human NK-2 receptor, we prepared a partial cDNA from human tracheal RNA using the polymerase chain reaction with oligonucleotide primers derived from the bovine NK-2 receptor cDNA sequence (Masu, Y., Nakayama, K., Tamaki, H., Harada, Y., Kuno, M., Nakanishi, S. (1987) Nature 329, 836-838). This partial human NK-2 receptor cDNA was used to screen a human genomic DNA library and yielded a clone, NGNK-2, of approximately 25 kilobases. Analysis of NGNK-2 indicates that it contains the entire coding sequence of the NK-2 receptor as well as 5'- and 3'-flanking sequences. The gene is organized with five exons interrupted by four introns. The complete sequence of the exons and the intron-exon junctions was determined, as were the transcription initiation site and the 3'-polyadenylation signal. Analysis of EcoRI digests of genomic DNA from human-mouse cell hybrids indicates a single gene for the human NK-2 receptor localized to chromosome 10. Sequence analysis of exons 1 and 5, where major differences occur between the human and animal species, provided information for polymerase chain reaction primers which allowed us to prepare full-length cDNA for the human NK-2 receptor. The protein predicted from the gene sequence is extended by 14 amino acids at the COOH terminus compared to the bovine and 9 residues compared to the rat molecules. The seven membrane-spanning regions are encoded by exons 1-4 and none is interrupted by introns. These regions are highly conserved among the species studied, suggesting stringent evolutionary control over these molecules.
Amino Acid Sequence↗
Structure and chromosomal location of the human gene encoding cartilage matrix protein.
Cartilage matrix protein (CMP) is a major component of the extracellular matrix of nonarticular cartilage. The structure and chromosomal location of the human gene encoding CMP was determined by molecular cloning analysis. We used a partial chicken CMP cDNA probe to isolate three overlapping human genomic clones. From one of these clones, a probe containing 2 human CMP exons was isolated and used to map the gene to chromosome 1p35 and to screen a human retina cDNA library. Two overlapping cDNA clones were isolated. The predicted protein sequence of 496 amino acids includes a 22-residue signal peptide and a 474-residue mature protein of Mr 51,344. The human CMP gene and polypeptide are strikingly similar to the chicken CMP gene and polypeptide. Human CMP is 79% identical to chicken CMP and contains two homologous domains separated by an epidermal growth factor-like domain. One potential N-glycosylation site is conserved between the two species. The human CMP gene spans 12 kilobase pairs with 8 exons and 7 introns which are similar in size to those of the chicken CMP gene. Both RNA splice junctions of intron G in the human and chicken CMP genes are nonconforming to the consensus splice sequences. This suggests that the CMP gene utilizes a new RNA splicing mechanism.
Amino Acid Sequence↗
Two chromosomal locations for human ornithine decarboxylase gene sequences and elevated expression in colorectal neoplasia.
The polyamines are known to be essential for cellular proliferation. Ornithine decarboxylase (ODC) is a rate-limiting enzyme in the synthesis of these amines, and activity is elevated in colorectal tumors and polyps. Two ODC genes (designated ODC1 and ODC2) were localized by somatic cell hybridization and in situ techniques to 2p25 and 7q31-qter, respectively. Investigation of the expression of ODC in colorectal neoplasia reveals a consistent increase in mRNA expression compared with normal adjacent mucosa and control mucosa, ranging from 1.3- to 12.2-fold. No amplification of the loci was seen. Comparison of ODC mRNA expression with ODC activity from the same samples revealed no direct correlation, suggesting that regulation of ODC in this system occurs at the posttranscriptional level.
Chromosome Mapping↗
Human facilitative glucose transporters. Isolation, functional characterization, and gene localization of cDNAs encoding an isoform (GLUT5) expressed in small intestine, kidney, muscle, and adipose tissue and an unusual glucose transporter pseudogene-like sequence (GLUT6).
Two novel facilitative glucose transporter-like cDNAs have been isolated from human small intestine and fetal skeletal muscle cDNA libraries by low stringency cross-hybridization with a fragment of the human erythrocyte/GLUT1 facilitative glucose transporter cDNA. One encodes a 501-amino acid facilitative glucose transporter, designated as the small intestine/GLUT5 isoform, having 41.7, 40.0, 38.7, and 41.6% identity with the previously described human erythrocyte/GLUT1, liver/GLUT2, brain/GLUT3, and muscle-fat/GLUT4 isoforms, respectively. GLUT5 mRNA is expressed at highest levels in small intestine and at much lower levels in kidney, skeletal muscle, and adipose tissue. Expression of in vitro synthesized human GLUT5 mRNA in Xenopus laevis oocytes indicates that the GLUT5 protein is a cytochalasin B-sensitive glucose carrier. The gene encoding the GLUT5 protein is located on the short arm of human chromosome 1. The second facilitative transporter-like cDNA sequence, designated GLUT6, is part of an 11-kilobase transcript that is expressed in all tissues examined. The sequence of a partial-length GLUT6 cDNA having an insert of 3.4 kilobase pairs revealed a region of 1.5 kilobase pairs that has 79.6% identity with the human brain/GLUT3 facilitative glucose transporter cDNA. However, because of the presence of multiple stop codons and frame shifts, this sequence cannot encode a functional glucose transporter protein. The region of facilitative glucose transporter nucleotide sequence homology in the GLUT6 transcript may have arisen by insertion of a reverse-transcribed GLUT3 transcript into the untranslated region of another gene. The GLUT6 gene is located on the long arm of human chromosome 5.
Adipose Tissue↗
Complete amino acid sequence of human cartilage link protein (CRTL1) deduced from cDNA clones and chromosomal assignment of the gene.
Little is known about the primary amino acid structure of human cartilage link protein (CRTL1). We screened a human genomic library with a cDNA encoding the 3' untranslated region and the adjoining B1 domain of chicken link protein. One clone was isolated and characterized. A 3.5-kb EcoRI-KpnI fragment from this genomic clone that contains the human B1 exon was used to map the gene to chromosome 5q13----q14.1. The same fragment was used to screen a cDNA library prepared from mRNA of Caco-2, a human colon tumor cell line. Two overlapping clones were isolated and shown to encode all of CRTL1. The deduced amino acid sequence is 354 residues long. The amino acid sequence shows a striking degree of identity to the porcine (96%), rat (96%), and chicken (85%) link protein sequences. Furthermore, there is greater than 86% homology between the 3' untranslated region of the genes encoding human and porcine link proteins. These results indicate that there has been strong evolutionary pressure against changes in the coding and 3' untranslated regions of the gene encoding cartilage link protein.
Amino Acid Sequence↗
Completion of the primary structure of the human type IV collagenase preproenzyme and assignment of the gene (CLG4) to the q21 region of chromosome 16.
The complete amino acid sequence of the human type IV collagenase preproenzyme was determined from cDNA and genomic clones. Primer extension and S1 nuclease analyses as well as nucleotide sequencing of a genomic clone indicate that the first exon has two closely spaced initiation sites for transcription and codes for 290 and 280 nt of a 5' untranslated region and a 29-residue signal peptide. The gene (CLG4) was localized to 16q21 using somatic cell hybrids and in situ hybridization.
Amino Acid Sequence↗