Search PubMed⌕ Search

Biomedical subjects

J Shine

Publications and source records attributed to J Shine.

At least 109 records · Page 6Linked to original sources
PubMed1984

Relaxin gene expression in human ovaries and the predicted structure of a human preprorelaxin by analysis of cDNA clones.

In earlier studies we identified in a human genomic library a gene (human relaxin gene H1) coding for a relaxin-related peptide. We now have evidence that the human genome possesses an additional relaxin-related gene (designated human relaxin gene H2) which appears to be selectively expressed in the ovary during pregnancy. Nucleotide sequence analysis revealed striking differences in the predicted structures of relaxin encoded by these two genes. Chemical synthesis of biologically active relaxin based on the sequence obtained from ovarian cDNA clones confirmed that the expressed gene (H2) encodes an authentic human relaxin. The expressed gene appears to be transcribed into two different sized mRNAs and preliminary evidence suggests that the mRNA transcripts possess different 3'-untranslated regions. There was no evidence for the expression of human relaxin gene H1 in the ovary and so far it is unclear whether gene H1 is expressed in another tissue or whether it represents a pseudogene. From the sequence data presented here it will now be possible to construct oligonucleotide probes and raise antibodies against synthetic peptides which could then be used to identify sites of relaxin biosynthesis and specifically quantitate the expression from either the H1 or H2 relaxin genes.

Amino Acid Sequence↗
PubMed1984

Two human relaxin genes are on chromosome 9.

We have recently cloned two different human relaxin gene sequences. One of these (H1) was isolated from a human genomic clone bank and the other (H2) from a cDNA library prepared from human pregnant ovarian tissue. Southern gel analysis of the relaxin genes within the genomes of several unrelated individuals showed that all genomes contained both relaxin genes. Hence it is unlikely (p less than 0.001) that the two relaxin gene sequences are alleles. Rather, it is probable that there are two relaxin genes within the human genome. It is likely that relaxin and insulin genes have evolved from a common ancestral gene by gene duplication, since structural similarities between insulin and relaxin are evident at both the peptide and gene level. To investigate the evolutionary relationship between the two human relaxin genes and the insulin gene, we have determined the chromosomal position of the relaxin genes using mouse/human cell hybrids. We found that the human insulin and relaxin genes are on different chromosomes. Both human relaxin genes are located on the short arm region of chromosome 9.

Alleles↗
PubMed1984

Primary structure of the human renin gene.

The gene encoding human renin has been isolated on two overlapping clones from a bacteriophage lambda library of human DNA. The entire gene spans about 12,000 bp and contains 10 exons separated by 9 intervening sequences. The gene structure is similar to that of human pepsinogen in terms of overall size, homology in the coding regions, position of introns, and sizes of the exons, suggesting that the two genes are evolutionarily related. However, a novel exon coding for only three amino acids was detected that is not present in the pepsinogen gene and whose amino acids are also not found in mouse renin. Although the nucleotide sequence of the 5'-flanking DNA differs from that of the pepsinogen gene, in both cases this region contains a structure of almost perfect dyad symmetry which immediately precedes the TATA box and may have functional importance. Furthermore, sequences resembling the putative consensus sequence for glucocorticoid regulation of gene expression are located approximately 200 and 300 bp upstream from the gene. The overall structural anatomy suggests that the human renin gene evolved by mechanisms that include a duplication of exon segments, particularly those containing the codons for the catalytically important aspartate residues, together with the insertion of other exon and flanking DNA structures. An analysis of human chromosomal DNA demonstrates that there is only one gene with high homology to human renin.

Base Sequence↗
PubMed1984

Human renin gene sequence, gene regulation and prorenin processing.

Human DNA coding for renin was identified and sequenced. The gene consisted of 10 exons corresponding to a 1500 nucleotide mRNA was broken up by long stretches of 'nonsense' DNA (introns) and spanned 12,000 base pairs. In addition, the sequence of nucleotides involved in regulation of the gene was determined by sequencing upstream. Prediction of the amino acid sequence of human preprorenin revealed likely sites of processing. This helps explain many past experimental observations. For example, the pro region contained adjacent likely cleavage sites for trypsin and pepsin and so reveals why both trypsin and pepsin can activate prorenin. The structure of human renin had features involved in its highly specific hydrolysis of the Leu10-Val11 bond unique to human angiotensinogen: in particular leucine 224 (instead of valine). Renin gene expression was studied in the mouse by quantification of both renin activity and its mRNA. Sodium depletion, captopril and spironolactone increased expression of Ren-1 in the kidney. The unusual, duplicated, mouse gene, Ren-2, which is expressed in the submandibular gland was, regulated by (dihydro)testosterone in male mice and by thyroid hormone in female mice.

Animals↗
PubMed1983

Structure of mouse kallikrein gene family suggests a role in specific processing of biologically active peptides.

The glandular kallikrein gene family comprises 25-30 highly homologous genes that encode specific proteases involved in the processing of biologically active peptides. In the mouse all the members of this family are closely linked on chromosome 7. The 9.5-kilobase nucleotide sequence of a mouse genomic clone contains one complete kallikrein gene (mGK-1), which is expressed in the male mouse submaxillary gland, and the 3' end of another (mGK-2). Differences in the coding potential of these genes and the amino acid sequences of other known kallikreins seem to be functionally related to the substrate specificity of the different enzymes.

Amino Acid Sequence↗
PubMed1983

In vitro expression of nitrogenase activity in Parasponia-Rhizobium strain ANU 289.

Rhizobium strain ANU 289 derepressed nitrogenase activity under defined in vitro conditions. Acetylene reduction was detected both in agar and liquid stationary culture. The strain is capable of nitrogen-fixing nodulation of legumes [such as siratro (Macroptilium atropurpureum Urb] as well as the non-legumes Parasponia andersonii and P. rugosa. Nitrogenase activity as high as 40-70 nmol C2H4 per mg protein after 7 days of incubation was detected. Strain ANU 289 was similar to Rhizobium strains 32 H1 and CB 756 with regard to oxygen requirement in the gas phase for development of nitrogenase activity between 0 and 10% O2, but showed increased sensitivity to oxygen repression at 20% O2. Strain ANU 289 also showed pronounced sensitivity to exogenous glutamine compared to strains 32 H1 and CB 756.

Nitrogenase↗
PubMed1983

Nitrogenase structural genes are unlinked in the nonlegume symbiont Parasponia rhizobium.

Several Rhizobium strains are capable of biological nitrogen fixation in symbiotic association with nonleguminous plants. The gene encoding the iron-protein component of nitrogenase (nifH) from one such strain, Parasponia Rhizobium sp. ANU289, has been isolated and completely sequenced. Unlike previously studied nitrogen-fixing prokaryotes, the Fe-protein subunit is encoded on a separate operon from other components of the nitrogenase enzyme complex. Comparative analysis of Fe-protein amino acid sequences indicates that the symbiotic nitrogen fixation phenotype in Rhizobium may have arisen on at least two separate occasions during its evolution.

Amino Acid Sequence↗
PubMed1983

Biological nitrogen fixation: primary structure of the Rhizobium trifolii iron protein gene.

Biological nitrogen fixation in the Rhizobium-legume symbiosis is dependent on the induction of a bacterially-encoded enzyme complex, nitrogenase. To examine the organization and expression of the genes encoding the components of nitrogenase in this complex system, these genes have been isolated from the legume symbiont Rhizobium trifolii by molecular cloning. DNA sequence analysis of the entire nifH gene (encoding the Fe-protein component of nitrogenase) and of the amino-terminal 141 codons of the nifD gene (encoding the alpha-subunit of the Mo-Fe protein) indicates that these genes are linked on a single operon in this strain. The Fe-protein amino acid sequence shares considerable homology with the sequence from other organisms, in particular the related organism Rhizobium meliloti (90% homology). The nif structural genes are preceded by a DNA sequence which is repeated at least three times in the Rhizobium trifolii genome.

Amino Acid Sequence↗
PubMed1983

Sym plasmid transfer to various symbiotic mutants of Rhizobium trifolii, R. leguminosarum, and R. meliloti.

Two self-transmissible Sym(biosis) plasmids, one encoding pea-specific nodulation and nitrogen-fixation functions (plasmid pJB5JI) and the other encoding clover-specific nodulation and nitrogen-fixation functions (plasmid pBR1AN) were used to determine whether the symbiotic genes encoded on these plasmids are expressed in various members of the Rhizobiaceae. The host specificity of Rhizobium trifolii and R. leguminosarum Sym plasmid-cured strains could be directly determined by the transfer to these strains of the appropriate Sym plasmid. The nodulation of white clovers was restored by either plasmid pJB5JI or pBR1AN when these plasmids were transferred to two transposon Tn5-induced hair-curling (Hac-) R. trifolii mutants. In addition, lucerne nodulation was restored to a Hac- R. meliloti mutant when either plasmid pBR1AN or pJB5JI was transferred to this strain. The phenotype of nonmucoid (Muc-) Rhizobium mutants, which had altered cell surfaces, was not influenced by the transfer to these strains of plasmid pBR1AN or plasmid pJB5JI.

Conjugation, Genetic↗
PubMed1983

Expression of the prolactin gene in human decidua-chorion.

Messenger RNA (mRNA) purified from human decidua-chorion hybridizes with a 32P-cDNA probe for human prolactin. In contrast, mRNA from human trophoblast and amnion does not hybridize to prolactin cDNA. The migration position on agarose gels of prolactin-specific mRNA from human decidua-chorion is similar to that for mRNA from ovine pituitary, suggesting a similar sized mRNA coding for prolactin in these different tissues and species. These data demonstrate unambiguously that the prolactin gene is expressed in human decidua-chorion, confirming previous immunochemical reports of prolactin production by this tissue in culture.

Amnion↗
PubMed1982

Mouse glandular kallikrein genes. Nucleotide sequence of cloned cDNA coding for a member of the kallikrein arginyl esteropeptidase group of serine proteases.

A library of cloned cDNA to male mouse submaxillary gland poly(A)-containing RNA was constructed in the plasmid pBR322. Inserts containing sequences estimated to be in the 1-5% abundance class were identified by hybridization to radiolabeled cDNA and examined by nucleotide sequence analysis. A sequence coding for a peptide with 57% homology to the only complete kallikrein sequence reported to date (from pig pancreas) was identified by a computer search program. This insert appears to code for the COOH-terminal 149 amino acids of a protein presumed therefore to be a serine protease. Comparison of the predicted amino acid sequence of this protein with analogous sequences in the three characterized members of the mouse submaxillary gland kallikrein arginyl esteropeptidase group of enzymes revealed extensive homology, although not complete identity. Thus, there are at least four members of this enzyme family expressed in the mouse submaxillary gland.

Amino Acid Sequence↗
PubMed1982

The human pro-opiomelanocortin gene: organization, sequence, and interspersion with repetitive DNA.

The human pro-opiomelanocortin (POMC) gene has been characterized by molecular cloning and DNA sequence analysis. Although this gene codes for several different polypeptide hormones, only a single intron interrupts the protein coding region. The DNA in this intron, and in a second intron found in the region of the gene homologous to the mRNA 5'-untranslated sequence, contains repetitive DNA sequences. At least some of these sequences belong to the Alu family of transcribed middle repetitive DNA. The determination of the complete nucleotide sequence of the coding regions of the gene demonstrates that the pattern of homologous and variable regions seen in the POMC protein between different species is reflected at the DNA level. DNA sequences encoding the highly conserved regions of POMC are 90-95% homologous between species while the coding sequences for the variable regions of the protein are approximately 70% homologous. The very high degree of homology in the amino terminal portion of POMC is consistent with an important physiological role for this peptide.

Amino Acid Sequence↗
PubMed1982

Porcine relaxin: molecular cloning and cDNA structure.

Relaxin is a peptide hormone produced by the corpora lutea of ovaries during pregnancy, softening and lengthening the ligaments of the pelvis and softening the cervix in order to make childbirth easier. In attempts to determine the nucleotide sequence coding for relaxin, recombinant DNA techniques were used to obtain a cDNA clone bank from total mRNA isolated from the ovaries of pigs in late pregnancy. Clones were screened using cDNA initiated by synthetic oligonucleotide primers coding for the Trp Val Glu Ile sequence of the porcine relaxin B chain. The synthetic undecamer [5'-ATCTCCACCCA-3'] was found to prime a specific 32P-labeled cDNA of approximately 300 nucleotides containing B chain and signal peptide coding sequences, as verified by nucleic acid sequence analysis. This cDNA was used to probe the ovarian clone bank. Several clones containing large inserts which hybridized to this probe were subjected to sequence analysis and some of these were found to contain the preprorelaxin coding region, comprising a signal peptide of 24 amino acids, a B chain of 32 amino acids, a large C peptide of 104 amino acids, and an A chain of 22 amino acids. From the amino acid sequence of prorelaxin derived in this way, it appears that the processing of prorelaxin involves two enzymes with chymotrypsin-like and trypsin-like specificity, respectively. In comparisons of porcine and rat preprorelaxins, the C region had as much amino acid sequence homology as the B and A chains. The C region is also rich in charged amino acids, suggesting a role for it beyond simply ensuring proper disulfide bond formation.

Animals↗
PubMed1982

Mouse kallikrein arginyl-esteropeptidase genes: analysis of cloned cDNAs suggests rapid functional divergence from a common ancestral sequence.

A previously-cloned cDNA coding for a member of the kallikrein arginyl-esteropeptidase group of serine proteases, (pMK-1), was used as a hybridization probe to identify a second partial cDNA clone (pMK-2) from mouse submaxillary gland. pMK-2 shares more than a 98% nucleotide sequence homology with pMK-1; the 3' untranslated regions are identical and there are only two predicted amino acid changes over the C-terminal 66 amino acids. The site of one change is implicated in determining substrate specificity, while the other may affect the catalytic mechanism. Thus despite the marked similarity of pMK-1 and pMK-2, the differences probably give rise to functionally distinct enzymes and are not simply polymorphic alleles.

Amino Acid Sequence↗