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C Gerald

Publications and source records attributed to C Gerald.

27 records · Page 2Linked to original sources
PubMed1995

Expression cloning and pharmacological characterization of a human hippocampal neuropeptide Y/peptide YY Y2 receptor subtype.

The pancreatic polypeptide family includes neuropeptide Y (NPY), one of the most abundant neuropeptides in the mammalian nervous system, as well as peptide YY (PYY) and pancreatic polypeptide (PP). This peptide family is involved in numerous physiological processes such as memory, pain, blood pressure, appetite, anxiety, and circadian rhythm. Of the multiple Y-type receptors proposed for PP family members, only the Y1 subtype was cloned previously. We now report the isolation of a human Y2 (hhY2) receptor cDNA by expression cloning from a human hippocampal cDNA library, using a 125I-PYY binding assay. hhY2 cDNA encodes a predicted protein of 381 amino acids with low amino acid identity to the human Y1 receptor (31% overall; 41% transmembrane). 125I-PYY binding to transiently expressed hY2 receptors was saturable (pKd = 10.17) and displaceable by human PP family members in rank order: PYY (pKi = 9.47) approximately NPY (pKi = 9.27) >> PP (pKi < 6) and by peptide analogs: NPY2-36 (pKi = 8.80) approximately NPY13-36 (pKi = 8.55) approximately C2-NPY (pKi = 8.54) > NPY26-36 (pKi = 6.51) approximately [Leu31,Pro34]NPY (pKi = 6.23). Human PYY decreased [cAMP] and increased intracellular [Ca2+] in hY2-transfected 293 cells.

Amino Acid Sequence↗
PubMed1995

The 5-HT4 receptor: molecular cloning and pharmacological characterization of two splice variants.

Molecular cloning efforts have provided primary amino acid sequence and signal transduction data for a large collection of serotonin receptor subtypes. These include five 5-HT1-like receptors, three 5-HT2 receptors, one 5-HT3 receptor, two 5-HT5 receptors, one 5-HT6 receptor and one 5-HT7 receptor. Molecular biological information on the 5-HT4 receptor is notably absent from this list. We now report the cloning of the pharmacologically defined 5-HT4 receptor. Using degenerate oligonucleotide primers, we identified a rat brain PCR fragment which encoded a '5-HT receptor-like' amino acid sequence. The corresponding full length cDNA was isolated from a rat brain cDNA library. Transiently expressed in COS-7 cells, this receptor stimulates adenylyl cyclase activity and is sensitive to the benzamide derivative cisapride. The response is also blocked by ICS-205930. Interestingly, we isolated two splice variants of the receptor, 5-HT4L and 5-HT4S, differing in the length and sequence of their C-termini. In rat brain, the 5-HT4S transcripts are restricted to the striatum, but the 5-HT4L transcripts are expressed throughout the brain, except in the cerebellum where it was barely detectable. In peripheral tissues, differential expression was also observed in the atrium of the heart where only the 5-HT4S isoform was detectable.

Adenylyl Cyclases↗
PubMed1994

Identification of a B2 bradykinin receptor expressed by PC12 pheochromocytoma cells.

We have used rat PC12 pheochromocytoma cells, a clonal cell line closely related to sympathetic neurons, to investigate reports that the bradykinin receptor expressed in the peripheral nervous system is distinct from the well-characterized B2 bradykinin receptor of smooth muscle. Although there have been reports that [Thi5,8,D-Phe7]bradykinin [where Thi is beta-(2-thienyl)alanine] is a full agonist at some sites in the peripheral nervous system, we find that in PC12 cells [Thi5,8,D-Phe7]bradykinin behaves as a competitive antagonist of bradykinin-stimulated phosphatidylinositol turnover. In particular, sufficient concentrations of [Thi5,8,D-Phe7]bradykinin completely block the increase in inositol bisphosphate and trisphosphate in response to 100 nM bradykinin; [Thi5,8,D-Phe7]bradykinin alone, at up to 10 microM, does not appreciably increase inositol bisphosphate and trisphosphate. In contrast to the absence of evidence for a distinctive neuronal receptor, we have found convincing evidence that the bradykinin receptor previously identified in smooth muscle is present in PC12 cells. Using the polymerase chain reaction, we have isolated a full-length cDNA encoding a bradykinin receptor that is expressed in PC12 cells and verified that its nucleotide sequence is identical except at a single position to that of the rat uterine B2 bradykinin receptor. When expressed in COS cells this uterine bradykinin receptor exhibits the same high affinity for [3H]bradykinin (Kd 4.4 nM), the same relative affinities for a series of kinin antagonists, and the same efficient coupling to phosphatidylinositol turnover (EC50 2.5 nM) as the receptor in PC12 cells. We interpret our data, and the findings of a number of pharmacological studies, as strengthening the view that the B2 receptor expressed in PC12 cells and in certain cells of the peripheral nervous system is identical to the receptor in rat uterine smooth muscle.

Adrenal Gland Neoplasms↗
PubMed1992

Molecular cloning of a complementary deoxyribonucleic acid encoding the thyrotropin-releasing hormone receptor and regulation of its messenger ribonucleic acid in rat GH cells.

Rat pituitary GH cells have been used extensively to study the biochemical actions of TRH on lactotropic cells. To investigate the structure and regulation of the rat TRH receptor (rTRHR), we have cloned its cDNA from GH4C1 cells. Using the polymerase chain reaction with degenerate primers and pools of cloned cDNAs from a GH4C1 cDNA library, a fragment sharing high similarity to the mouse thyrotrope TRHR (mTRHR) was identified. Conventional library screening with this fragment was used to isolate a single cDNA. mRNA synthesized in vitro from this cDNA was injected into Xenopus oocytes, and a characteristic conductance response to TRH was detected by voltage clamp recording. DNA sequence analysis revealed a molecule of 412 amino acid residues, with 96% similarity to the mTRHR. However, in contrast to the mTRHR, the rTRHR had an additional 19 amino acid residues at its carboxy-terminus. A mRNA of about 4 kilobases was identified in GH3 cells. Regulation of the rTRHR mRNA concentration was studied in GH3 cells. Steady state rTRHR mRNA levels were decreased to 30% of the control level by incubation with TRH for 48 h and increased 4-fold by incubation with dexamethasone for 12 h. Southern blot analysis of genomic DNA from GH3 cells gave a simple banding pattern consistent with a single copy gene. We conclude that the rTRHR shares high primary sequence similarity to the mTRHR, but the rTRHR has an extension of 19 amino acids at its carboxy-terminus, which is lacking in the mTRHR.

Amino Acid Sequence↗
PubMed1987

Fusion glycoprotein of measles virus: nucleotide sequence of the gene and comparison with other paramyxoviruses.

The sequence of the fusion (F) glycoprotein mRNA of the Hallé strain of measles virus was determined from a cDNA clone representing the entire length of the mRNA. It contained 2384 nucleotides, excluding poly(A), with a 5' consensus sequence typical of paramyxoviruses and a 3' terminus found in measles virus mRNAs. The coding sequence was preceded by an unusually long (580 nucleotide) 5' non-translated region, which contained 44% cytosine. The longest open reading frame coded for a polypeptide of 553 amino acids with a predicted molecular weight of 59.84 K. Comparison of the sequence with that of the Edmonston strain of measles virus showed that the gene is highly conserved. No amino acid differences were observed between the two strains. The F polypeptide had three regions of high hydrophobicity: an N-terminal signal peptide, the N-terminus of F1 and a C-terminal membrane-spanning region. The four potential asparagine-linked glycosylation sites (one in the signal peptide) were all in the F2 subunit. Comparison of the measles virus F amino acid sequence with other paramyxoviruses revealed homologies with these viruses. Certain regions such as the N terminus of F1 and ten cysteine residues which probably impose structural restraints were highly conserved.

Amino Acid Sequence↗
PubMed1986

Measles virus haemagglutinin gene: cloning, complete nucleotide sequence analysis and expression in COS cells.

A measles virus (Hallé strain) cDNA library was prepared by cloning virus-induced mRNA directly into the expression vector PCD. Clones corresponding to the measles virus haemagglutinin (HA) gene were isolated and one, PCD-HA-15, which corresponded to the complete mRNA sequence, was further characterized. After transfection into COS-7 cells, measles virus HA antigen was detected by immunofluorescence. The [35S]methionine-labelled HA protein from transfected cells was immunoprecipitated by both polyclonal and monoclonal measles virus antibodies. Analysis by SDS-polyacrylamide gel electrophoresis revealed that the PCD-HA-15 protein migrated in a manner identical to the virus-induced HA. Nucleotide sequence analysis established that the gene contained 1949 nucleotides [exclusive of poly(A)] and coded for a protein containing 617 amino acids. A single hydrophobic domain likely to represent the transmembrane region was identified at the N-terminus. A second overlapping reading frame coded for a protein containing 70 amino acids. This contained a short hydrophobic region (16 amino acids) and had two potential N-glycosylation sites. Comparison of the HA gene of the Hallé strain with the published sequence of the Edmonston strain showed that there was a high degree of conservation (99.3%).

Amino Acid Sequence↗
PubMed1984

A study of measles virus antigens in acutely and persistently infected cells using monoclonal antibodies: differences in the accumulation of certain viral proteins.

Monoclonal antibodies were used to study measles virus (Hallé strain) infections. The distribution of virus antigens in acutely and persistently infected cells was examined by immunofluorescence, and the synthesis of the virus polypeptides in acutely infected cells was measured after immunoprecipitation and analysis in SDS-polyacrylamide gels. The virus nucleoprotein (NP) was detected 12 h after infection and was the major virus polypeptide synthesized throughout the virus cycle. The NP, matrix, and polymerase (L) polypeptides were first detected associated with the perinuclear membrane, but were later distributed throughout the cytoplasm. The hemagglutinin accumulated in the cell membrane in well-defined 'clusters'. In comparison with the acutely infected system, persistently infected cells presented important differences. There were only low levels of L polypeptides, and up to 50% of the cells contained intranuclear inclusions which stained specifically with NP monoclonal antibody. The relevance of these observations is discussed.

Animals↗