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H M Kronenberg

Publications and source records attributed to H M Kronenberg.

At least 109 records · Page 6Linked to original sources
PubMed1985

Messenger ribonucleic acid from tumors associated with humoral hypercalcemia of malignancy directs the synthesis of a secretory parathyroid hormone-like peptide.

Previous studies have provided evidence that tumors associated with humoral hypercalcemia of malignancy produce a factor that shares certain biological properties with, but is not, native PTH. In the present study, media from Xenopus oocytes microinjected with polyadenylated RNA prepared from three human or animal tumors associated with humoral hypercalcemia of malignancy are shown to have activity in the cytochemical bioassay for PTH. This activity parallels that of the PTH standard in the assay and is completely inhibited by the PTH analog [Nle8,Nle18,Tyr34]bovine PTH-(3-34)NH2. Media from oocytes injected with mRNA from control tumors contain no such activity. The mRNA encoding this activity in one of the animal tumors has been enriched approximately 10-fold by preparative polyacrylamide gel electrophoresis. These results demonstrate that tumors associated with humoral hypercalcemia of malignancy have the capacity for directing the synthesis of a secretory protein that fulfills certain criteria for being the humoral mediator in question. These techniques may provide the basis for molecular cloning of this factor.

Animals↗
PubMed1984

Gene encoding parathyroid hormone. Nucleotide sequence of the rat gene and deduced amino acid sequence of rat preproparathyroid hormone.

The nucleotide sequence of the rat parathyroid hormone (PTH) gene was established from a 14.5-kilobase pair fragment of rat liver DNA cloned in bacteriophage Charon 4A. The transcriptional unit of the gene of 2.5 kilobase pairs is interrupted by two introns (1600 and 111 base pairs). Blot hybridization of restriction enzyme digests of rat spleen DNA using 32P-labeled fragments of the cloned PTH gene suggests that the gene is unique and present in a single copy in the genome. A promoter sequence (Goldberg-Hogness or TATA box) is situated 28 base pairs upstream from the point of initiation of transcription which was found by S1 nuclease mapping and by oligonucleotide-primed reverse transcription of rat PTH mRNA. The gene is flanked on its 5' side by repetitive DNA and contains a different, more abundant repetitive DNA on its 3' side. The mRNA encoded by the parathyroid hormone gene consists of 800 +/- 50 nucleotides as determined by electrophoresis on agarose gels. The 5' untranslated region of the mRNA contains three AUG triplets. Only one AUG triplet initiates biosynthesis of preproparathyroid hormone (prepro-PTH). The 3' untranslated segment of the mRNA contains two AATAAA sequences characteristic of polyadenylation signals. The mRNA encodes prepro-PTH, a precursor of PTH of 115 amino acids. When the amino acid sequence of the rat precursor is compared with the analagous bovine and human precursors, it becomes evident that the hormone sequences are highly conserved in two regions of known function near the NH2 terminus and in a third region near the carboxyl terminus whose biologic function, if any, has not yet been defined.

Amino Acid Sequence↗
PubMed1984

Secretion of human parathyroid hormone from rat pituitary cells infected with a recombinant retrovirus encoding preproparathyroid hormone.

In order to study the functions of precursors to secreted proteins, we expressed cloned DNA encoding human preproparathyroid hormone (preproPTH) in rat pituitary cells. We first constructed a recombinant plasmid containing human preproPTH cDNA and retroviral control signals. This recombinant plasmid was transfected into psi-2 cells, a packaging cell line that produces Moloney murine leukemia viral particles containing no retroviral RNA. The transfected psi-2 cells generated helper-free recombinant retrovirus encoding preproPTH, and this recombinant retrovirus was used to infect GH4 rat pituitary cells. Clonal lines of the infected GH4 cells contained copies of the recombinant provirus stably integrated via the long terminal repeats, and the expected RNA transcripts of proviral DNA accumulated in the cytoplasm, although no infectious virus was produced. The infected cells synthesized and processed preproPTH appropriately and secreted PTH in response to thyrotropin-releasing hormone, a secretagogue for GH4 cells. Use of recombinant retrovirus permits the introduction of DNA encoding normal and mutant secreted proteins into a number of cell types specialized for secretion. Analysis of the fate of the resultant proteins will help define the specific molecular interactions involved in transmembrane transport and processing of precursor proteins.

Amino Acid Sequence↗
PubMed1983

Human parathyroid hormone gene (PTH) is on short arm of chromosome 11.

The human gene for parathyroid hormone (PTH) was chromosomally mapped using human-rodent hybrids and Southern filter hybridization of cell hybrid DNA. A recombinant DNA probe containing human PTH cDNA insert (pPTHm122) hybridized to a 3.7-kb fragment in human DNA cleaved with the restriction enzyme EcoRI. By correlating the presence of this fragment in somatic cell hybrid DNA with the human chromosomal content of the hybrid cells, the PTH gene was mapped to the short arm of the chromosome 11.

Animals↗
PubMed1983

Beta-globin locus is linked to the parathyroid hormone (PTH) locus and lies between the insulin and PTH loci in man.

Using a parathyroid hormone (PTH) cDNA probe we found a common Pst I polymorphic restriction site 3' to the PTH gene in all ethnic groups examined. Because the PTH, insulin, and beta-globin loci have been localized to the short arm of chromosome 11 (11p) we used DNA polymorphisms adjacent to each of these three loci to determine whether they are genetically linked and to determine their order. We found that the PTH and beta-globin loci are closely linked (estimated recombination fraction, 0.07; 95% confidence limits, 0.05-0.10; lod score, 4.63; odds favoring linkage, 42,000:1). Furthermore, our findings strongly indicate that the beta-globin gene cluster lies between the PTH and insulin loci. Therefore, the gene order on 11p is centromere-PTH-beta-globin-insulin.

Chromosomes, Human, 6-12 and X↗
PubMed1983

Nucleotide sequence of the human parathyroid hormone gene.

From a lambda phage gene library we have isolated phage containing the gene encoding human preproparathyroid hormone. The phage were isolated by using both the plaque-hybridization technique and the in vivo recombination-selection technique. The human preproparathyroid hormone gene contains two intervening sequences that separate the gene into a 5' noncoding domain, a "prepro" sequence domain, and a domain containing the parathyroid hormone sequence and the 3' noncoding region. The gene is approximately 4,200 base pairs long. Restriction endonuclease analysis of human leukocyte DNA shows that the haploid human genome contains one copy of the preproparathyroid hormone gene. A 14-base-pair sequence of alternating purines and pyrimidines that has the potential of adopting the Z-DNA conformation lies 134 base pairs upstream from the presumed site of initiation of transcription.

Amino Acid Sequence↗
PubMed1983

Transport and cleavage of bacterial pre-beta-lactamase by mammalian microsomes.

Eucaryotic endoplasmic reticulum and bacterial inner cell membranes use strikingly similar mechanisms to transport proteins from one cellular compartment to another. Using a mammalian linked transcription-translation system, we show here that canine pancreatic microsomes accurately cleave off the signal sequence of beta-lactamase, a secreted bacterial protein. We suggest that the apparent differences between bacterial and eukaryotic protein transport may involve only minor modulations of a profoundly conserved mechanism.

Animals↗
PubMed1983

Parathyroid hormone messenger ribonucleic acid: effects of calcium on cellular regulation in vitro.

The effects of changes in the concentration of extracellular calcium the principal regulatory of PTH secretion, on the cellular levels of the mRNA encoding prepro-PTH, a precursor of PTH, were studied in bovine parathyroid slices in vitro using a sensitive radiodensitometric cDNA hybridization assay and mRNA translation in a wheat germ cell-free system. Although lowering the calcium from 5 to 0.5 mM during 5- to 70-h incubations stimulated the secretion of PTH 4- to 8-fold and pro-PTH synthesis 1.4-fold, the cellular levels of total hybridizable PTH mRNA did not change. However, a 2- to 3-fold increase in PTH mRNA, which could be hybridized to oligo-dT-cellulose [poly(A)-rich], was observed in the slices incubated in 5 mM calcium (suppressed slices). The poly(A)-rich PTH mRNA constituted 30% of the total PTH mRNA in suppressed gland slices and 10% of the total PTH mRNA in stimulated gland slices. PTH mRNA did not decay measurably when RNA synthesis in the gland slices was inhibited with actinomycin D and alpha-amanitin for 6 h. Both the stability and the lack of perturbation of the cellular levels of total of total hybridizable PTH mRNA by changes in the concentration of extracellular calcium suggest that the short term regulation of PTH biosynthesis involves posttranscriptional mechanisms.

Animals↗
PubMed1982

Looking at genes.

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Anemia, Sickle Cell↗
PubMed1981

Nucleotide sequence of cloned cDNAs encoding human preproparathyroid hormone.

We have cloned cDNA copies of human preproparathyroid hormone in Escherichia coli after insertion of double-stranded DNA into the Pst I site of plasmid pBR322 using the poly(dG) . poly(dC) homopolymer extension technique. Recombinant plasmids coding for preproparathyroid hormone were identified by filter hybridization assay using as a probe 32P-labeled bovine preproparathyroid cDNA. Nucleotide sequence analysis of five recombinant plasmids permitted the assignment of 74 nucleotides of the 5' noncoding region, the entire coding region of 345 nucleotides, and the entire 3' noncoding region of 348 nucleotides of the mRNA. The coding sequence predicts the previously unknown "pre" amino acid sequence and clarifies the hormone's amino acid sequence, which has been disrupted. The 5' noncoding region contains an AUG codon followed by a UGA stop codon before the authentic initiator codon. The 3' noncoding region is 120 nucleotides longer than in bovine preproparathyroid mRNA and contains two A-A-U-A-A-A sequences, potential signals for polyadenylation.

Base Sequence↗
PubMed1981

Nucleotide sequence of the mRNA encoding the pre-alpha-subunit of mouse thyrotropin.

We have constructed and cloned in bacteria recombinant DNA molecules containing DNA sequences coding for the precursor of the alpha subunit of thyrotropin (pre-TSH-alpha). Double-stranded DNA complementary to total poly(A)+RNA derived from a mouse pituitary thyrotropic tumor was prepared enzymatically, inserted into the Pst I site of the plasmid pBR322 by using poly(dC).poly(dG) homopolymeric extensions, and cloned in Escherichia coli chi 1776. Cloned cDNAs encoding pre-TSH-alpha were identified by their hybridization to pre-TSH-alpha mRNA as determined by cell-free translations of hybrid-selected and hybrid-arrested RNA. The nucleotide sequences of two cDNAs (510 and 480 base pairs) were determined with chemical methods and corresponded to much of the region coding for the alpha subunit and the 3' untranslated region of pre-TSH-alpha mRNA. The sequence of the 5' end of the mRNA was determined from cDNA synthesized by using total mRNA as template and a restriction enzyme DNA fragment as primer. Together these sequences represented greater than 90% of the coding and noncoding regions of full-length pre-TSH-alpha mRNA, which was determined to be 800 bases long. The amino acid sequence of the pre-TSH-alpha deduced from the nucleotide sequence showed a NH2-terminal leader sequence of 24 amino acids followed by the 96-amino-acid sequence of the apoprotein of TSH-alpha. There is greater than 90% homology in the amino acid sequences among the murine, ruminant, and porcine alpha subunits and 75-80% homology among the murine, equine, and human alpha subunits. Several regions of the sequence remain absolutely conserved among all species, suggesting that these particular regions are essential for the biological function of the subunit. The successful cloning of the alpha subunit of TSH will permit further studies of the organization of the genes coding for the glycoprotein hormone subunits and the regulation of their expression.

Animals↗
PubMed1980

Biosynthesis of parathyroid hormone.

Figure 11 summarizes our present concepts of the biosynthetic sequence for parathyroid hormone, deduced largely from observations in vitro. Many aspects of the presumed process whereby preproparathyroid hormone is converted via proparathyroid hormone to the hormone remain unclarified and require much further study, as is true for many other prehormones. These studies, however, coupled with (1) further investigations of intracellular degradation of parathyroid hormone, if this indeed operates in vivo; (2) the proteolytic conversion of secreted hormone in peripheral tissues; and (3) analysis of transcriptional control of biosynthesis of parathyroid hormone, using radioactive cDNA for hybridization studies of mRNA production and turnover, hold great promise for further understanding of critical regulatory factors central to expression of the actions of parathyroid hormone.

Adenylyl Cyclases↗
PubMed1979

Cloning and nucleotide sequence of DNA coding for bovine preproparathyroid hormone.

We have cloned in Escherichia coli a DNA copy of mRNA coding for bovine preproparathyroid hormone. Double-stranded DNA was inserted into the Pst I site in plasmid pBR322 by using the poly(dG)-poly(dC) homopolymer extension technique to join the DNA molecules. Recombinant plasmids coding for preproparathyroid hormone were identified by the plasmid's ability to arrest specifically the translation of preproparathyroid hormone mRNA. The nucleotide sequence of the largest recombinant was determined by using both chemical and enzymatic techniques. The parathyroid insert contains 470 nucleotides--102 nucleotides from the 5' noncoding region of the mRNA, 345 nucleotides representing the entire coding region, and 23 nucleotides from the 3' noncoding region. The coding sequence clarifies the hormone's amino acid sequence, which has been disputed. Codon usage is discussed.

Amino Acid Sequence↗
PubMed1978

Pre-proparathyroid hormone; amino acid sequence, chemical synthesis, and some biological studies of the precursor region.

The precursor of bovine proparathyroid hormone was synthesized by translation of parathyroid mRNA in a wheat-germ cell-free system. The amino acid sequence of the NH2-terminal extension (the pre sequence) was determined by repetitive Edman degradation of the polypeptide labeled with radioactive amino acids (radiosequencing). The pre sequence of pre-proparathyroid hormone is (formula: see text) which is followed by the sequence of proparathyroid hormone. It is significant that 20 of the 25 amino acids in the sequence are hydrophobic. This high hydrophobicity is consistent with the proposed role of the pre sequence as a membrane-penetrating peptide. The precursor-specific sequence of 31 amino acids was snythesized chemically by the solid-phase technique. This synthetic peptide was shown to bind to the microsomal fraction of homogenates prepared from extracts of parathyroid glands, a finding consistent with the proposed role of the precursor peptide in the attachment of the nascent chain--mRNA--ribosome complex to the endoplasmic reticulum.

Amino Acid Sequence↗