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J Ham

Publications and source records attributed to J Ham.

At least 73 records · Page 4Linked to original sources
PubMed1987

Identification of androgen response elements in mouse mammary tumour virus and the rat prostate C3 gene.

The effect of steroid hormones on mouse mammary tumour virus (MMTV) promoter activity has been analysed in the breast cancer cell lines ZR-75-1 and T-47D 11. Both androgens and progestins were stimulatory in both cell lines, whereas glucocorticoids were stimulatory in only ZR-75 cells. A glucocorticoid response was restored to T-47D cells by the transient expression of a glucocorticoid receptor expression vector, suggesting that endogenous receptors were limiting. The effect of androgen was mediated by a region of MMTV previously shown to be essential for glucocorticoid and progestin responses between nucleotides -201 and -69 upstream of the cap site. Similarly, the effect of androgens on the rat prostate C3(1) gene promoter was analysed in the rat prostate cell line T5. Although these cells had functional androgen receptors which could stimulate MMTV-promoter activity, they were incapable of interacting with androgen response elements to stimulate transcription from the C3(1) promoter.

Androgens↗
PubMed1987

Structural organization and expression of the mouse estrogen receptor.

Complementary DNA clones corresponding to the mouse uterus estrogen receptor mRNA have been isolated and characterized. Nucleotide sequence analysis predicts that full-length cDNA has the potential to code for a polypeptide of 599 amino acids, and comparison with the protein sequences of the rat, human, and chicken estrogen receptors reveals overall homologies of 97%, 88% and 77%, respectively. Genomic clones for the mouse estrogen receptor have been isolated from a cosmid library and used in conjunction with the cDNA clones to study the expression of the receptor in vivo by RNase mapping, primer extension, and Northern blotting. These analyses demonstrate that transcription initiates at multiple sites which span a region of at least 62 base pairs and that the estrogen receptor is encoded by mRNA of approximately 6.5 kilobases in size. There are 10 major starts in total, one of which is situated 31 nucleotides downstream from a TATA box-like motif and coincides with the start of the cDNA clone pMOR8. The ability of the cDNA clone to produce a functional protein was verified by transfection into COS-1 cells which lack endogenous estrogen receptor. The mouse estrogen receptor, in a SV40-based expression vector, was cotransfected with a chimeric marker plasmid consisting of an estrogen response element from the vitellogenin A2 gene linked to the thymidine kinase promoter and the chloramphenicol acetyl transferase gene. In the presence of estradiol chloramphenicol acetyl transferase activity is stimulated by up to 80-fold, while tamoxifen and 4-hydroxytamoxifen act primarily as antiestrogens in this in vitro assay.

Amino Acid Sequence↗
PubMed1986

Chronic stimulation of anterior pituitary cell cultures with CRF leads to the secretion of lipotropin.

Time course studies have shown that the secretion of beta-endorphin-related peptides from primary cultures of rat anterior pituitary reached a plateau after 1 h of stimulation by 10(-7) M corticotropin-releasing factor (CRF); the major peptide released was beta-endorphin 1-31. Further incubation of the cells with CRF led to a decline in the rate of secretion and to the appearance of a different pattern of peptides in which lipotropin was the principal component; analysis of the peptides remaining in the cells, however, showed that beta-endorphin was still the major peptide. Chronic blockade of the anterior pituitary cells with dexamethasone (10(-6) M) resulted in diminished secretion and the released peptides were found to contain a higher proportion of the intact prohormone, pro-opiomelanocortin (POMC). In all the experiments involving stimulation of the pituitary cells with CRF or blockade with corticosteroids it was observed that the pattern of secreted peptides differed from the intracellular patterns, which suggests that selective release can occur from vesicles or cells that contain different segments of POMC.

Animals↗
PubMed1985

beta-Endorphin and ACTH related peptides in primary cultures of rat anterior pituitary cells: evidence for different intracellular pools.

Acid extracts of rat anterior pituitary cells and cell-derived culture media were shown to contain three forms of beta-endorphin immunoreactive peptides, corresponding in molecular size to the prohormone pro-opiomelanocortin (POMC), beta-lipotropin and 3.5 kDa beta-endorphin, and essentially two forms of adrenocorticotropin (ACTH) immunoreactivity, representing a 20 kDa intermediate fragment and 4.5 kDa ACTH. Under basal conditions the intracellular peptides contained a high proportion of the bioactive forms of beta-endorphin and ACTH whereas the extracellular peptides contained a higher proportion of the inactive precursors. When the cells were incubated for 3 h in the presence of 10(-8) M CRF, the levels of intracellular beta-endorphin and ACTH immunoreactivity were reduced by 15-30% and there was a 4-5 fold increase in the level of the secreted peptides; furthermore, unlike the peptides released under basal conditions, the peptides secreted under the influence of CRF contained much higher proportions of 4.5 kDa ACTH and 3.5 kDa beta-endorphin, reflecting the intracellular patterns of these peptides. Similar results were obtained when secretion was stimulated by 10(-7) M epinephrine, which produced a 2-fold increase in peptide release. In the presence of 10(-6) M dexamethasone the basal secretion of ACTH and beta-endorphin related peptides, and the intracellular levels of these peptides, remained unaltered. The results point to the existence of different intracellular compartments from which peptides at different states of maturation can be released selectively.

Adrenocorticotropic Hormone↗
PubMed1985

Beta-endorphin processing in pituitary and brain is sensitive to haloperidol stimulation.

Chronic administration of haloperidol (1 mg/Kg) into rats led to a marked increase in the level of beta-endorphin related peptides in the pars intermedia and brain stem. Using a combination of ion-exchange chromatography and radioimmunoassay, the increase in beta-endorphin under haloperidol treatment was confined to the acetylated derivatives: alpha,N-acetyl beta-endorphin 1-26, alpha,N-acetyl beta-endorphin 1-27 and alpha,N-acetyl beta-endorphin 1-31; there was no change in the level of the NH2-peptides. Haloperidol had no effect on the beta-endorphin related peptides in the anterior pituitary or hypothalamus. These findings suggest that haloperidol can influence not only the level of beta-endorphin related peptides in pituitary and brain, but also their posttranslational processing, possibly serving to regulate the level of biologically active beta-endorphin 1-31.

Animals↗
PubMed1985

Cloning of the CDC7 gene of Saccharomyces cerevisiae in association with centromeric DNA.

The cell-division-cycle gene, CDC7, of Saccharomyces cerevisiae has been cloned in a large plasmid (pCM6) containing an insert of about 45 kb of yeast chromosomal DNA in the yeast-Escherichia coli shuttle vector, YRp7. A subclone (pCM39), having a 7-kb insert with a unique BamHI site, was capable of conferring the ts+ phenotype on a cdc7ts- strain of S. cerevisiae. When this insert was placed in the vector Y1p28, and then cut with BamHI and integrated into the ts- strain, genetic analysis showed the CDC7-complementing activity and an associated vector marker to be linked to the TRP1 locus, i.e., to chromosome IV. Analysis of subclones covering the pCM39 insert showed that pCM52, in which a 3.3-kb EcoRI-HpaI fragment with the unique BamHI site has been cloned, possessed the full complementing activity. Although the CDC7 gene lies across the BamHI site, the main part of the gene appears to be in the 2.2-kb BamHI-HpaI segment of the pCM52 insert. A Northern blot analysis showed this region to give rise to a 1.7-kb mRNA which we conclude is the transcript of CDC7. The large insert of pCM6 also contains a 1.5-kb XhoI fragment which several observations suggest is that containing the centromere (CEN4) of chromosome IV to which the CDC7 gene is closely linked. The 45-kb DNA fragment in pCM6 is believed to be an uninterrupted piece of chromosome IV.

Cell Cycle↗
PubMed1984

Regulation of bioactive beta-endorphin processing in rat pars intermedia.

Acid extracts of rat pituitary neuro-intermediate lobes have been shown by ion-exchange chromatography and radio-immunoassay to contain predominantly the inactive derivatives of beta-endorphin, alpha, N-acetyl beta-endorphin 1-27 and alpha, N-acetyl beta-endorphin 1-26; the biologically active form, beta-endorphin 1-31, is a minor component. In contrast, it was found that beta-endorphin generated in neuro-intermediate lobe cells in monolayer culture was less processed: the principal peptides related to bioactive beta-endorphin 1-31. When the cultured cells were incubated in the presence of 10(-5) M dopamine or 10(-6) M alpha-ergocryptine there was a marked increase in the degree of proteolysis and acetylation: the processing pattern reverted to that characteristic of the neuro-intermediate lobe in situ, with alpha-N-acetyl beta-endorphin 1-26 and alpha, N-acetyl beta-endorphin 1-27 as the prominent peptides. The results demonstrate that dopaminergic agents can influence the processing of beta-endorphin-related peptides in rat pars intermedia, indicating a new level at which the bioactivity may be regulated.

Animals↗
PubMed1984

The use of high-performance liquid chromatography in the isolation and characterisation of mouse and rat epidermal growth factors and examination of apparent heterogeneity.

Various epidermal growth factor preparations obtained from the mouse submaxillary gland (mEGF), have been separated into a number of components by reversed-phase high-performance liquid chromatography (HPLC). It is shown here, however, that when the mEGF is isolated rapidly, using only reversed-phase HPLC for trace enrichment and high-resolution fractionation, it is a single molecular species as determined with several ion-pairing solvent systems, provided that proteolysis is inhibited in the original extracts. This indicates that the minor components of mEGF that have been reported are artefacts formed during the isolation procedure, and are of no biological significance. The products of deliberate mild degradation of mEGF are shown to produce similar chromatographic profiles to those observed in samples of mEGF prepared in the absence of proteolytic inhibitors. Rat EGF has been isolated in a similar manner, and is shown to share many of the properties of the major tryptic digestion product of mEGF.

Amino Acids↗
PubMed1980

Purification and partial characterization of high-molecular-weight forms of ectopic calcitonin from a human bronchial carcinoma cell line.

Studies on the high-molecular-weight immunoreactive calcitonin produced ectopically in culture by an epidermoid bronchial carcinoma cell line are reported. In cell-exposed medium, the principal component has a molecular weight of 40000 and molecules of mol.wts. 13000 and 10000 also occur. Only a trace amount of material co-eluting with 35000-mol.wt. human calcitonin is detectable. None of the calcitonins show cross-reactivity with anti-corticotropin serum. The 40000-mol.wt. immunoreactive calcitonin is readily proteolysed to the 13000- and 10000-mol.wt. components, but the 10000-mol.wt. component behaves as a comparatively stable 'core' molecule. By using immunoprecipitation and high-pressure liquid chromatography (h.p.l.c.), it is possible to prepare radiochemically homogeneous 10000-mol.wt. immunoreactive calcitonin from cells grown in the presence of individual 35S- or 3H-labelled amino acids. Peptide mapping of enzymic digests of this material by h.p.l.c. shows that it contains peptides in common with synthetic human calcitonin.

Adenocarcinoma, Bronchiolo-Alveolar↗
PubMed1980

Tumour calcitonin. Interaction with specific calcitonin receptors.

The human epidermoid bronchial carcinoma (BEN) cell line has been shown to have specific membrane binding sites for calcitonin and to secrete high-molecular-weight forms (ranging from 40000 to 10000) of immunoreactive calcitonin. Synthetic salmon and human calcitonins and a thyroid extract of porcine calcitonin have been shown to displace 125I-labelled salmon calcitonin from the receptors in a dose-related fashion. The binding to these receptors of calcitonins derived from the BEN cell line and a medullary thyroid carcinoma with molecular weights ranging from 28000 to 3500 (both separated by gel-filtration chromatography) has been investigated. Neither major peaks of BEN-cell-line calcitonin showed receptor binding activity. Only one form of medullary thyroid carcinoma calcitonin, that which co-eluted with synthetic calcitonin monomer on gel-filtration chromatography, caused any significant displacement of labelled hormone from the receptors.

Animals↗
PubMed1979

Simultaneous modeling of pharmacokinetics and pharmacodynamics: application to d-tubocurarine.

We propose a model of drug pharmacodynamic response that when integrated with a pharmacokinetic model allows characterization of the temporal aspects of pharmacodynamics as well as the time-independent sensitivity component. The total model can accommodate extremes of effect. It allows fitting of simultaneous plasma concentration (Cp) and effect data from the initial distribution phase of drug administration, or from any non-equilibrium phase. The model postulates a hypothetical effect compartment, the dynamics of which are adjusted to reflect the temporal dynamics of drug effect. The effect compartment is modeled as an additional compartment linked to the plasma compartment by a first-order process, but whose exponential does not enter into the pharmacokinetic solution for the mass of drug in the body. The hypothetical amount of drug in the effect compartment is then related to the observed effect by the Hill equation, a nonlinear sigmoid form. Nonlinear least-squares data fitting is used for parameter estimation. The model is demonstrated on two different sets of Cp and effect data for the drug d-tubocurarine (dTC). In 7 normal subjects, the (mean +/- SD) rate constant for equilibration of dTC effect (paralysis) and Cp is 0.13 +/- 0.04 min-1 and the (mean +/- SD) steady-state Cp required to produce 50% paralysis is 0.37 +/- 0.05 microgram/ml.

Adult↗