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R L Eddy

Publications and source records attributed to R L Eddy.

At least 37 records · Page 2Linked to original sources
PubMed1995

The Brn-3 family of POU-domain factors: primary structure, binding specificity, and expression in subsets of retinal ganglion cells and somatosensory neurons.

A search for POU domain sequences expressed in the human retina has led to the identification of three closely related genes: Brn-3a, Brn-3b, and Brn-3c. The structure and expression pattern of Brn-3b was reported earlier (Xiang et al., 1993); we report here the structures and expression patterns of Brn-3a and Brn-3c. Antibodies specific for each Brn-3 protein were generated and shown to label only ganglion cells in a variety of vertebrate retinas. A complex pattern of strongly and weakly immunolabeled ganglion cells was observed in mouse, cat, and monkey retinae. In mouse and cat retinae, Brn-3a and Brn-3b proteins are found in a large fraction of ganglion cells, whereas Brn-3c is present in fewer ganglion cells. In the cat retina, anti-Brn-3a immunoreactivity was strong in the small ganglion cells (gamma cells) and weak in the remaining ganglion cells (alpha and beta cells); anti-Brn-3b immunoreactivity was present in all ganglion cells; and anti-Brn3c immunoreactivity was confined to the small ganglion cells. Immunolabeling of macaque retinae following retrograde labeling from the lateral geniculate nucleus revealed strong anti-Brn-3a immunoreactivity in a minority of retrogradely labeled P-type ganglion cells, and weak Brn-3a immunoreactivity in all of the remaining P- and M-type ganglion cells. In the same retinae, strong anti-Brn-3b immunoreactivity was seen in nearly all P-type ganglion cells and weak immunoreactivity in nearly all M-type ganglion cells. Each of the Brn-3-specific antibodies also labeled subsets of neurons in the dorsal root and trigeminal ganglia, suggesting that primary somatosensory neurons and retinal ganglion cells share genetic regulatory hierarchies. In vitro selection of an optimal DNA binding site using the Brn-3b POU domain has revealed a consensus [(A/G)CTCATTAA(T/C)] that is recognized by each of the Brn-3 POU domains and is distinct from binding sites previously described for other POU domain proteins.

Amino Acid Sequence↗
PubMed1994

Type I procollagen COOH-terminal proteinase enhancer protein: identification, primary structure, and chromosomal localization of the cognate human gene (PCOLCE).

Type I procollagen COOH-terminal proteinase (C-proteinase) enhancer, a glycoprotein that binds to the COOH-terminal propeptide of type I procollagen and enhances procollagen C-proteinase activity, was purified from mouse fibroblast culture media. Partial amino acid sequences obtained from proteolytic fragments were found to have identity with the deduced amino acid sequence of a cDNA clone of unknown function, previously isolated from a mouse astrocyte library. Sequences of mouse enhancer cDNA, obtained in the present study, predict a approximately 50-kDa, 468-amino acid protein that differs from the 43-kDa, 402-amino acid protein predicted by the previously reported astrocyte-derived clone. Human cDNAs encode an enhancer of 449 amino acids. Previous biochemical studies have found the mouse enhancer as a 55-kDa form, which is readily processed to 36- and 34-kDa forms, retaining full C-proteinase enhancing activity and the ability to bind the COOH-terminal propeptide. Data presented here show the 36-kDa form to correspond to the amino-terminal portion of the 55-kDa protein. This is the most conserved region between mouse and human enhancers, comprising two domains with homology to domains found in a number of proteases and proteins with developmental functions. Such domains are thought to mediate interactions between proteins. Mouse enhancer RNA is shown to be at highest levels in collagen-rich tissues, especially tendon. The human enhancer gene, PCOLCE, is localized to 7q21.3-->q22, the same chromosomal region containing the type I collagen alpha 2 chain gene, COL1A2.

Amino Acid Sequence↗
PubMed1994

Human, mouse, and rat calnexin cDNA cloning: identification of potential calcium binding motifs and gene localization to human chromosome 5.

Calnexin is a 90-kDa integral membrane protein of the endoplasmic reticulum (ER). Calnexin binds Ca2+ and may function as a chaperone in the transition of proteins from the ER to the outer cellular membrane. We have purified human calnexin in association with the human interferon-gamma receptor and cloned calnexin cDNA from placenta. Fragments of calnexin have been prepared as glutathione S-transferase fusion proteins and analyzed for their abilities to bind 45Ca2+ and ruthenium red. A subdomain containing four internal repeats binds Ca2+ with the highest affinity. This sequence is highly conserved when compared to calreticulin (a luminal ER protein), an Onchocerca surface antigen, and yeast and plant calnexin homologues. Consequently, this sequence represents a conserved motif for the high-affinity binding of Ca2+, which is clearly distinct from the "E-F hand" motif. An adjacent subdomain, also highly conserved and containing four internal repeats, fails to bind Ca2+. The carboxyl-terminal, cytosolic domain is highly charged and binds Ca2+ with moderate affinity, presumably by electrostatic interactions. The calnexin amino-terminal domain (residues 1-253) also binds Ca2+, in contrast to the amino-terminal domain of calreticulin, which is relatively less acidic. We have also determined the cDNA sequences of mouse and rat calnexins. Comparison of the known mammalian calnexin sequences reveals very high conservation of sequence identity (93-98%), suggesting that calnexin performs important cellular functions. The gene for human calnexin is located on the distal end of the long arm of human chromosome 5, at 5q35.

Amino Acid Sequence↗
PubMed1994

Cloning of human and mouse EBI1, a lymphoid-specific G-protein-coupled receptor encoded on human chromosome 17q12-q21.2.

A lymphoid-specific member of the G-protein-coupled receptor family has been identified by PCR with degenerate oligonucleotides. We have determined that this receptor, also reported as the Epstein-Barr-induced cDNA EBI1, is expressed in normal lymphoid tissues and in several B- and T-lymphocyte cell lines. While the function and the ligand for EBI1 remain unknown, its sequence and gene structure suggest that it is related to the receptors that recognize chemoattractants, such as interleukin-8, RANTES, C5a, and fMet-Leu-Phe. Like the chemoattractant receptors, EBI1 contains intervening sequences near its 5' end; however, EBI1 is unique in that both of its introns interrupt the coding region of the first extracellular domain. The gene is encoded on human chromosome 17q12-q21.2. None of the other G-protein-coupled receptors has been mapped to this region, but the C-C chemokine family has been mapped to 17q11-q21. The mouse EBI1 cDNA has also been isolated and encodes a protein with 86% identity to the human homolog.

Amino Acid Sequence↗
PubMed1993

Structural organization and chromosomal assignment of the gene encoding the human heparin-binding epidermal growth factor-like growth factor/diphtheria toxin receptor.

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a recently identified, potent smooth muscle cell mitogen of macrophage origin. It is expressed in a highly regulated fashion in vascular endothelial and smooth muscle cells, indicating a potentially important role for this gene in atherosclerosis. In addition, the HB-EGF precursor has recently been found to function as a receptor for diphtheria toxin. Using an HB-EGF cDNA probe, we cloned the human gene encoding HB-EGF. The HB-EGF gene contains six exons and five intervening sequences spanning 14 kb of DNA. By primer extension and S1 nuclease analysis, we located a major transcription start site (corresponding to an A residue) 14 bp beyond the 5' end of the HB-EGF cDNA. There were no TATAAA or CCAAT consensus sequences upstream of the transcription start site. The density of primer extension bands generated by RNA from endothelial cells treated with tumor necrosis factor-alpha (TNF-alpha) was 10 times higher than that of bands generated by the control, indicating that TNF-alpha increased the level of HB-EGF mRNA. Using transient reporter gene transfection experiments, we show that 2.0 kb of HB-EGF 5'-flanking sequence has promoter activity in bovine aortic endothelial cells. By analysis of DNA isolated from human-mouse somatic hybrid cell lines, we assign the HB-EGF gene to chromosome 5. By functional study, chromosome 5 has been associated with diphtheria toxin susceptibility.

Amino Acid Sequence↗
PubMed1993

Chromosome mapping and organization of the human beta-galactoside alpha 2,6-sialyltransferase gene. Differential and cell-type specific usage of upstream exon sequences in B-lymphoblastoid cells.

The human beta-galactoside alpha 2,6-sialyltransferase (EC 2.4.99.1) (SiaT-1) gene is localized to human chromosome 3 (q21-q28) by Southern analysis of somatic cell hybrids and by in situ hybridization of metaphase chromosomes. Comparative analysis between the human and the previously reported rat SiaT-1 genomic sequences demonstrates precise conservation of the intron/exon boundaries throughout the coding domains. Furthermore, there is extensive inter-species sequence similarity in some of the exons that contain information only for the 5'-leader regions. Human genomic sequences were also analyzed to reconcile reported differences in the 5'-untranslated region in SiaT-1 mRNAs. In cultured cell lines of the B-lineage, Reh, Nalm-6, Jok-1, Ball-1, Daudi, and Louckes, the study demonstrates that three upstream exons, Exons(Y+Z) and Exon(X), are mutually exclusively utilized, resulting in at least two distinct populations of SiaT-1 mRNA being synthesized. None of these exons is present in the SiaT-1 mRNA isotype expressed in HepG2 human hepatoma cells. In all B-lymphoblastoid cell lines examined, the basal level SiaT-1 mRNA is maintained by the expression of an isotype containing the Exons(Y+Z) sequence. The slightly smaller SiaT-1 mRNA, which contains the Exon(X) sequence but not Exons(Y+Z) sequence, is synthesized at a high level and found only in Jok-1, Daudi, and Louckes, the cell lines with mature B-cell phenotype. The study also provides further evidence that induced SiaT-1 expression accompanies the appearance of CDw75, a putatively sialylated cell surface epitope and a marker of human mature B-lymphocytes. The SiaT-1 induction is the result of the appearance of a novel form of SiaT-1 mRNA isotype.

Animals↗
PubMed1993

Human chemotaxis receptor genes cluster at 19q13.3-13.4. Characterization of the human C5a receptor gene.

The human C5a anaphylatoxin and formyl peptide receptor genes, as well as two genes with high sequence identity to the formyl peptide receptor, FPRH1 and FPRH2, have been mapped to chromosome 19 (Lu et al., 1992). Further analysis reveals that these genes are present in the 19q13.3 band adjacent to the 13.3-13.4 interface. MRNAs for the C5a and formyl peptide receptors, as well as for FPRH1, are expressed in cAMP differentiated U937 cells and human eosinophils, while all four transcripts are expressed in human lung. This observation opens the possibility for coordinate regulation of these genes. In order to initiate the mapping of fine structure at this locus, genomic clones have been analyzed. All four of the genes have a similar structure, with the receptor protein encoded in a single exon. Detailed characterization of the C5a receptor gene reveals a two exon structure, with the 5' untranslated sequence and initiating methionine located in the first exon. An intron of approximately 9 kb separates exon 1 from the receptor-encoding exon 2. The region of genomic DNA flanking the 5' untranslated sequence possesses promoter activity when transfected into the myeloid-derived rat basophilic leukemia RBL-1 cells, but the same region is inactive when transfected into nonmyeloid cells. Deletional analyses indicate that C5a receptor 5' flanking region contains both cell-specific suppressor and promoter regions.

Animals↗
PubMed1993

Molecular analysis of a t(11;14)(q23;q11) from a patient with null-cell acute lymphoblastic leukemia.

/lp;&-3qChromosome 11, band q23, is the frequent site of recurring cytogenetic rearrangements in human leukemia. We have cloned and sequenced the breakpoint junctions from a patient who had null-cell acute lymphoblastic leukemia (ALL) with a t(11;14)(q23;q11). The chromosome 14 breakpoints occurred within the TCRD locus, close to two diversity segments. The chromosome 11 breakpoint occurred between two head-to-head heptamer sequences, and junctional diversity was evident at both derivative junctions, suggesting involvement of the V(D)J recombinase. The TCRA/D locus on the normal chromosome 14 had undergone a V delta 2-D delta 3-psi J alpha joining. Two phage clones with this VDJ rearrangement were isolated; one of these contained an intra-J alpha region deletion. Two clones with the derivative 11 junction were isolated; one of these had a similar, but not identical, deletion. A heptamer-nonamer recognition sequence (located approximately 70 kb 5' to C alpha), not associated with a TCR gene coding segment, was found in the immediate vicinity of both 5' breakpoints. We have designated this sequence 5'del for 5' deleting element. An intra-J alpha region deletion involving this heptamer-nonamer was previously identified in the leukemia cells recovered from a patient who had T-cell ALL. Fifty kilobases of DNA on 11q23 surrounding the breakpoint were cloned and analyzed. No CpG islands or conserved sequences were identified within this region.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

The genes for the lipopolysaccharide binding protein (LBP) and the bactericidal permeability increasing protein (BPI) are encoded in the same region of human chromosome 20.

The lipopolysaccharide binding protein is an acute-phase reactant produced during gram-negative bacterial infections. The bactericidal/permeability increasing protein is associated with human neutrophil granules and has bactericidal activity on gram-negative organisms. In addition to their functional relationship, both proteins share extensive structural similarity. This article demonstrates that the genes for both proteins are in the same region of human chromosome 20, between q11.23 and q12.

Acute-Phase Proteins↗
PubMed1993

Cloning of a portion of the chromosomal gene and cDNA for human beta-fodrin, the nonerythroid form of beta-spectrin.

A 96-bp synthetic oligonucleotide corresponding to an amino acid sequence near the N-terminus of erythroid beta-spectrin was used to screen a human genomic library, and two overlapping recombinants were isolated. DNA sequence analysis established that the genomic fragment encoded beta-fodrin, the nonerythroid form of beta-spectrin, by correlation to a known amino acid sequence of human brain beta-fodrin. The genomic DNA contained regions that cross-hybridized with an erythroid beta-spectrin cDNA probe, and the DNA sequence of these regions revealed a high degree of identity with that of erythroid beta-spectrin and a similar exon/intron organization. A single-copy DNA fragment of the beta-fodrin genomic clone was used to screen a lymphoid cell cDNA library and two recombinants were isolated. The composite DNA sequence of these various genomic and cDNA clones encoded almost all of the first twelve 106 amino acid repeat segments of beta-fodrin that shared 58% identity and 75.5% similarity with the amino acid sequence of beta-spectrin and 66% identity with the nucleotide sequence of beta-spectrin cDNA. The chromosomal localization of the gene was determined to be chromosome 2 by hybridization of a single-copy probe derived from the cloned genomic DNA to DNA of a panel of somatic hybrid cell lines, and in situ hybridization localized the gene to band 2p21. beta-Fodrin was assigned the gene symbol SPTBN1.

Amino Acid Sequence↗
PubMed1993

Linkage of Agt and Actsk-1 to distal mouse chromosome 8 loci: a new conserved linkage.

Angiotensinogen is an alpha 2-globulin involved in the maintenance of blood pressure and electrolyte balance. We have refined the position of the mouse angiotensinogen locus (Agt) on Chromosome (Chr) 8 and have also confirmed the assignment of the human angiotensinogen locus (AGT) to Chr 1. The segregation of several restriction fragment length variants (RFLVs) was followed in two interspecific backcross sets and in four recombinant inbred (RI) mouse sets. Analysis of the segregation patterns closely linked Agt to Aprt and Emv-2, which places the angiotensinogen locus on the distal end of mouse Chr 8. Additionally, a literature search has revealed that the strain distribution pattern (SDP) for the mouse skeletal alpha-actin locus 1 (Actsk-1, previously Acta1, Acta, or Acts) is nearly identical to the SDP for Agt in two RI sets. On the basis of this information we were able to reassign Actsk-1 to mouse Chr 8. By screening a panel of human-mouse somatic cell hybrids, we confirmed that the human angiotensinogen locus lies on Chr 1. This information describes a new region of conserved linkage homology between mouse Chr 8 and human Chr 1. It also defines the end of a large region of conserved linkage homology between mouse Chr 8 and human Chr 16.

Actins↗
PubMed1993

Mapping of the genes for human endoplasmic reticular heat shock protein gp96/grp94.

The murine tumor rejection antigen gp96 (TRA1, mapped to mouse chromosome 10) is a member of the heat shock protein family. Using a fragment of the murine gp96 cDNA as a probe, three gp96-related human genes have been isolated and structurally characterized. They have been mapped to human chromosomes 1 (p22), 12 (q24.2-->q24.3), and 15 (q25-->q26) by Southern blot hybridization and in situ hybridization of gene-specific probes. Only one of the genes, designated TRA1 (human chromosome 12) is a coding gene; the other genes (TRA1P1 and TRAP2) appear to be independently derived, processed pseudogenes.

Animals↗
PubMed1993

Brn-3b: a POU domain gene expressed in a subset of retinal ganglion cells.

A search for POU domain transcription factors in human retina cDNA has led to the identification of Brn-3b, a class IV POU domain protein. Immunohistochemical experiments show that chicken, mouse, rabbit, monkey, and human retinas contain Brn-3b exclusively within a subpopulation of ganglion cells. In the adult mouse brain, Brn-3b is found only within cells in the deep layers of the superior colliculus, in the dorsal periaqueductal gray, and in a small cluster of cells in the brain stem near the area postrema. During the immediate postnatal period, cells containing Brn-3b are distributed in a number of regions within the brain stem and cerebellum. These data suggest that Brn-3b plays a role in determining and/or maintaining the identities of a small number of neurons, including a subset of visual system neurons.

Amino Acid Sequence↗
PubMed1993

Localization of the human collagen gene COL7A1 to 3p21.3 by fluorescence in situ hybridization.

An 8-kb genomic probe, containing 34 collagen-encoding exons, was localized to 3p21.3 by fluorescence in situ hybridization. The genomic probe encoded a previously uncharacterized carboxyl terminal portion of the alpha 1(VII) collagen chain. This mapping result confirms the previous assignment of the alpha 1(VII) gene (COL7A1) to 3p21 and offers a finer subregional localization than was previously available.

Animals↗
PubMed1993

Chromosomal localization and nucleoside diphosphate kinase activity of human metastasis-suppressor genes NM23-1 and NM23-2.

Human metastasis-suppressor genes nm23-1 (NME1) and nm23-2 (NME2) are implicated in control of the metastatic potential of malignant cells. Using somatic cell hybrid analysis and fluorescence in situ hybridization we co-localized both genes to 17q21.3. The 17q21 region carries the locus responsible for early-onset familial breast-ovarian cancer and several other genes that are involved in tumorigenesis and differentiation and undergo frequent rearrangements during neoplastic development. Thus, our mapping places the NME genes in a region that may be subjected to multiple selection pressures. NME1 and NME2 genes were expressed as soluble proteins in a T7 bacterial expression system. Both proteins are independently active nucleotide diphosphate kinases and readily form intra- and intermolecular disulfide bonds. The biochemical properties of these proteins may explain the diversity of mature eucaryotic nucleoside diphosphate kinases.

Animals↗
PubMed1992

A novel gene constitutively expressed in human lymphoid cells is inducible with interferon-gamma in myeloid cells.

A cluster of at least six interferon-gamma (IFN gamma)-inducible genes designated Ifi201-204 and located on mouse chromosome 1 has recently been described. Here, we report a human IFN-gamma-inducible gene, IFI 16, which has nucleotide sequence similarity with portions of two of the mouse genes, Ifi202 and Ifi204. A full-length cDNA clone derived from IFI 16 [2.709 kilobases (kb)] contained a single open reading frame of 2.187 kb which encoded a putative polypeptide of 729 amino acids and a predicted non-glycosylated M(r) of 80020. IFI 16 mRNA was found to be constitutively expressed in lymphoid cells and in cell lines of both the T and B lineages. By contrast, the mRNA was not expressed by the cell lines HL-60, U937, and K562, which represent early stages of myeloid development, but was strongly inducible in HL-60 and U937 with IFN-gamma. The IFI 16 protein demonstrated a putative domain structure with patchy similarity to the proteins expressed from genes Ifi202 and Ifi204. The mouse and human proteins each contain two analogous approximately 200 amino acid domains which are imperfect copies, but IFI 16 demonstrated additional unique regions, including a Lys-rich N-terminal portion and a "spacer" region between the reiterated domains, analogous to spacer regions in the CD5 and CD8 alpha molecules. Using a panel of inter-species somatic cell hybrid cell lines, IFI 16 was localized to the chromosomal region 1q12----1qter, a region syntenic between mouse and man. DNA blotting indicated that, in contrast to the mouse, IFI 16 is present as a single copy gene in the human genome.

Amino Acid Sequence↗
PubMed1992

Genomic organization and chromosomal location of the human gene encoding the B-lymphocyte activation antigen B7.

The human B lymphocyte activation antigen B7 provides regulatory signals for T lymphocytes as a consequence of binding to its ligands CD28 and CTLA-4. The cDNA for B7 has previously been isolated and predicted to encode a type I membrane protein. The predicted polypeptide has a secretory signal peptide followed by two contiguous Ig-like domains, a hydrophobic transmembrane region and a short cytoplasmic tail. Here we report the exon-intron genomic organization of human B7 and the chromosomal location. The gene has six exons that span approximately 32 kilobases of DNA. Exon 1 is not translated and the second exon contains the initiation ATG codon and encodes a predicted signal peptide. This gene structure is characteristic for several eukaryotic genes with tissue-specific expression. The third and fourth exons correspond to two Ig-like domains whereas the fifth and sixth exons encode respectively the trans-membrane portion and the cytoplasmic tail. This close relationship between exons and functional domains is a characteristic feature of genes of the Ig superfamily. Cell surface expression of the B7 gene product has previously been mapped to human chromosome 12 by antibody reactivity with the B7-specific monoclonal antibody BB-1. We here demonstrate that the B7 gene is located to the q21-qter region of chromosome 3 by DNA blot analysis of human x rodent somatic cell hybrids.

Amino Acid Sequence↗