Biomedical subjects

H G Friesen

Publications and source records attributed to H G Friesen.

At least 73 records · Page 4Linked to original sources

PubMed1989

G-proteins modulate prolactin- and interleukin-2-stimulated mitogenesis in rat Nb2 lymphoma cells.

ADP ribosylation in the presence of cholera or pertussis toxin indicated the presence of G-proteins in Nb2 cell membranes. Two protein bands, with mol wt of 43.5K and 46.5K, were radiolabeled by cholera toxin, while a single protein (41.5K mol wt) was ADP ribosylated by pertussis toxin. Northern hybridization of total RNA from Nb2 cells with specific cDNA probes indicated the presence of mRNA transcripts encoding Gs, Gi2, Go, and, to a lesser extent, Gi3. A characteristic of receptors coupled to G-proteins is that their binding properties are regulated by guanine nucleotides. The binding of [125I]human GH to the lactogen receptor as well as the binding of [125I]IL-2 to the IL-2 receptor were decreased in a dose-dependent manner by GTP, GDP, and the analog guanosine 5'-O-(3-thiotriphosphate). GMP, however, had no effect. The addition of pyruvate kinase and phosphoenolpyruvate to regenerate GTP from GDP greatly increased the apparent potency of GTP. Cholera toxin inhibited PRL- and interleukin-2-stimulated DNA synthesis and cell proliferation in the Nb2 cells. In contrast, pertussis toxin had a differential effect on PRL- and IL-2-stimulated cells. Pertussis toxin, at an optimal concentration of 0.01 ng/ml, significantly enhanced the stimulatory effects of PRL on DNA synthesis (P less than or equal to 0.01; n = 9) and cell proliferation (P less than or equal to 0.05; n = 9) compared with the effect of PRL alone. However, at higher concentrations the toxin inhibited PRL-stimulated DNA synthesis and cell proliferation. Complete inhibition was achieved with 1000 ng/ml toxin. In contrast to the biphasic effect on PRL-stimulated cells, pertussis toxin was only weakly inhibitory to cells treated with IL-2. At the highest concentration tested, pertussis toxin (1000 ng/ml) inhibited IL-2-stimulated DNA synthesis and cell growth by only 30-35%. (Bu)2cAMP (IC50 = 0.019 mM) or methylxanthine (MIX; IC50 = 0.25 mM) also inhibited PRL-stimulated DNA synthesis. In the absence of mitogen, neither agent, from 0.0001-1 mM, had any effect on DNA synthesis. Similarly, IL-2-stimulated DNA synthesis in Nb2 cells was inhibited by (Bu)2cAMP (IC50 = 0.019 mM) or MIX (IC50 = 0.072 mM). However, MIX was approximately 3 times as potent in inhibiting the cell response to IL-2 as that to PRL. The susceptibility of Nb2 cells to both bacterial toxins suggests a role for G-proteins in regulating PRL- or IL-2-stimulated mitogenesis in these cells.(ABSTRACT TRUNCATED AT 400 WORDS)

1-Methyl-3-isobutylxanthine↗

PubMed1989

Cleaved prolactin and its 16K fragment are generated by an acid protease.

A cleaved form of rat PRL (cldPRL), which resolves after reduction of the disulfide bonds into two fragments of 16K and 8K mol wt, has been reported in the pituitary gland. To further understand the genesis of this cleaved PRL and its related 16K fragment (cldPRL/16K), pituitary proteins were labeled with [35S]methionine and then dialyzed and lyophilized before sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Pulse-chase experiments did not show a precursor-product relationship between PRL and cldPRL. However, if proteins were concentrated by trichloroacetic acid instead of dialysis, no cldPRL/16K was observed. Other experiments demonstrated that during dialysis cldPRL/16K is generated in vitro at the expense of intact PRL in a time-dependent manner. Labeled proteins dialyzed against water previously adjusted to different pHs showed after electrophoresis that cldPRL/16K was generated only at acidic pH. In the presence of the protease inhibitors phenylmethylsulfonylfluoride, tosylamidelysyl chloromethylketone, or tosylphenylalanine chloromethylketone or when the incubation medium was heated at 65 C for 45 min, the acid pH was unable to generate cldPRL/16K. Trypsin inhibitors did not prevent cldPRL generation. In conclusion, an acid protease, possibly of lysosomal origin, cleaves PRL in vitro. cldPRL/16K form was not found in the pituitary or incubation medium under physiological conditions. This PRL variant is generated as a preparative artifact unless experimental conditions are controlled.

Animals↗

PubMed1989

Regulation of prolactin secretion in the human B-lymphoblastoid cell line IM-9-P3 by dexamethasone but not other regulators of pituitary prolactin secretion.

We examined whether the ectopic production of human(h) PRL by the human B-lymphoblastoid cell line IM-9-P3 is under hormonal control. We demonstrate that PRL secretion in this cell line is regulated by dexamethasone but not by other hormones known to modulate PRL secretion in the pituitary or decidua. Dexamethasone caused a reduction of secretion rates to 30% of control values after 3 day paralleled by a decrease in hPRL mRNA levels, without affecting cell viability. Half-life determinations of hPRL mRNA in control and dexamethasone-treated cells revealed a reduction of t1/2 from 16 to 4 h. PRL secreted by IM-9-P3 cells did not control its own secretion in an autocrine loop, nor did it serve as an autocrine growth factor.

Antibodies, Monoclonal↗

PubMed1989

Elevated expression of basic fibroblast growth factor messenger ribonucleic acid in acoustic neuromas.

Human tumors were analyzed for the presence of mRNA coding for basic fibroblast growth factor (basic FGF). Basic FGF transcript levels were consistently elevated in schwannoma samples (five acoustic neuromas and two spinal schwannomas) ranging from 9- to 22-fold higher than the average level of expression in four benign meningioma samples. Acidic extracts of acoustic neuromas contained a potent mitogen which bound to heparin-Sepharose, eluted at 2 M NaCl, and cross-reacted with an N-terminal specific anti-basic FGF antiserum. The present findings indicate that basic FGF appears to be the major heparin-binding endothelial cell mitogen in acoustic neuromas. Southern restriction analysis revealed no evidence of amplification or rearrangement of the gene for basic FGF in schwannomas or in the astrocytoma cell line U87-MG. These findings demonstrate a tumor-specific elevation in basic FGF transcript levels in tumors of Schwann cell origin and suggest that increased transcription or stabilization of basic FGF mRNA may play an autocrine role in the development and progression of these tumors.

Blotting, Southern↗

PubMed1989

Fibroblast growth factor release by bovine endothelial cells and human astrocytoma cells in culture is density dependent.

Basic fibroblast growth factor (bFGF) is a potent endothelial cell mitogen whose actions are mediated by binding to specific cell surface receptors on a variety of cell types. However, the amino acid sequence of bFGF does not contain a classical signal peptide sequence and the extent to which cellular stores of this mitogen are released is still a matter of some controversy. In the present study we examined the release of immunoreactive bFGF into serum-free conditioned medium of bovine corneal endothelial cells (BCE) and a human astrocytoma cell line, U87-MG. Western blotting analysis of BCE conditioned medium using N-terminal specific anti-bFGF serum revealed a single immunoreactive band of 32 kilodaltons, which was reduced to 18 kilodaltons in the presence of 8 M urea. Using a sensitive two-site immunoradiometric assay we were able to quantify the release of immunoreactive bFGF into the culture medium by BCE cells and by the human astrocytoma cell line U87-MG. In each case the release of bFGF was cell density dependent, but under all conditions the level of bFGF released was significantly greater in the transformed astrocytoma line, ranging from 15- to 50-fold higher than in the BCE cultures under various conditions. At 30% confluence the concentration of immunoreactive bFGF in the medium was maintained at a constant level for up to 24 h. However, the level of immunoreactive bFGF declined rapidly in confluent cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1989

An immunological cryo-ultrastructural study of a sequential appearance of proteins in placental binucleate cells in early pregnancy in the cow.

Using the most sensitive immunocytochemical method available, on ultrathin frozen sections, the results in this paper demonstrate that bovine placental lactogen (bPL) is present in the earliest fetal binucleate cells found at 21 days post coitum in the trophectoderm. A second protein, the SBU-3 antigen, which is absent in the early stages of pregnancy appears abruptly in the binucleate cell granules at 30 days post coitum coincident with the start of villus development. Subsequently, the granules contain both bPL and the SBU-3 antigen. This sequential production of unlike proteins indicates that the binucleate cell has different functions depending on the stage of pregnancy and has important roles to play both at implantation and in villus development.

Animals↗

PubMed1988

Comparative biochemistry and physiology of teleocalcin from sockeye and coho salmon.

This is a comparative study of the glycoprotein hormone, teleocalcin, from the corpuscles of Stannius of sockeye (Oncorhynchus nerka) and coho (O. kisutch) salmon. Coho teleocalcin was purified by the same procedures used previously to obtain sockeye teleocalcin and was obtained in a comparable yield. Both salmon teleocalcins had the same molecular weight as estimated by sodium dodecyl sulfate-electrophoresis and both appeared to have the structure of disulfide-linked oligomers. The two hormones were similar on the basis of amino acid and carbohydrate composition and shared 95% homology in the first 40 residues on the N-terminal. The salmon teleocalcins also shared 80% homology with the predicted 1-40 N-terminal sequence from Australian eels (Anguilla australis). Both teleocalcins had potent inhibitory effects on gill calcium uptake in intact rainbow trout (Salmo gairdneri). However, these effects were observed only at the peak in the calcium uptake cycle that is displayed by this species. In North American eels (A. rostrata), the acute administration of both teleocalcins caused significant inhibition of gill calcium uptake without any concomitant changes in plasma calcium levels or other plasma electrolytes. In 4- and 7-day stanniectomy (STX) eels, the acute administration of coho teleocalcin significantly reduced or completely abolished the accelerated gill calcium transport that occurs postoperatively, with no concomitant changes in plasma electrolytes or post-STX hypercalcemia. These experiments provide further evidence that teleocalcin is a regulator of gill calcium transport and has no acute hypocalcemic effects in fish.

Amino Acid Sequence↗

PubMed1988

Stimulation of growth of Nb2 lymphoma cells by interleukin-2 in serum-free and serum-containing media.

The human recombinant alanine-125 analogue of interleukin-2 (IL-2) causes a dose-dependent mitogenic response in rat lymphoma Nb2-11C cloned cells when tested in serum-containing medium and serum-free medium. IL-2 and hGH elicit their growth stimulation through different receptors since IL-2 does not compete with 125I-hGH for binding to Nb2 cells and Met14hGH, an antagonist of hGH, inhibits the hGH-stimulated growth of Nb2 cells but not that caused by IL-2. The tumor promoter, 12-O-tetradecanoyl phorbol-13-acetate (TPA) potentiates the action of hGH on Nb2 cells grown in both serum-containing and in serum-free medium. TPA to a variable degree also potentiates IL-2-stimulated growth of Nb2 cells when cultured in medium containing serum but has no effect on cells grown in serum-free medium. In conclusion, IL-2 is a potent mitogen for Nb2-11C cells.

Animals↗

PubMed1988

Relationship between bioassay and radioimmunoassay measurement of prolactin in the IPL rat, a hypoprolactinemic rat strain.

IPL (Institut Pasteur, Lyon) nude, hypoprolactinemic rats exhibit delayed puberty and a complete lack of lactation. To characterize the secretion of circulating forms of prolactin (PRL) of these rats, PRL concentrations were measured in serum and pituitaries of males and females under various physiological conditions. Two assay methods, a radioimmunoassay (RIA) and a sensitive bioassay (NB2BA) were employed. Normal rats of the Sprague-Dawley strain were tested simultaneously, as controls. The pituitary content of PRL, estimated either by RIA or by NB2BA, in IPL nude males and females was similar to that of normal male and female rats. On the contrary, serum PRL levels of IPL male rats, measured by RIA or NB2BA, were significantly reduced when compared to normal rats. In both groups, there was a close correlation between the results obtained by the two methods, the NB2BA estimates being higher. However, the NB2BA/RIA ratio was significantly decreased in serum from IPL nude rats compared to controls, indicating that the circulating form of PRL was less bioactive in this group. Castrated male rats injected with estradiol showed sharply increased PRL values as estimated by RIA or NB2BA. The increase was greater (35-fold) in IPL nude rats then in normal rats (9-fold), but these increases resulted in serum PRL levels being similar in the two groups. However, the NB2BA/RIA ratio remained significantly reduced in IPL nude rats. In female rats, PRL was measured during different physiological states: estrus, diestrus, proestrus at 1000, 1200, and 1600 h and Days 1 and 21 of gestation and 2 days postpartum.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1988

Differential effects of estrogen and growth hormone on uterine and hepatic insulin-like growth factor I gene expression in the ovariectomized hypophysectomized rat.

The acute and chronic effects of 17 beta-estradiol (E2) and GH on uterine and hepatic insulin-like growth factor I (IGF-I) gene expression in ovariectomized hypophysectomized (ovx-hypox) rats were examined. Six hours after a single injection of E2 (5 micrograms/100 g BW), uterine IGF-I gene expression was increased 22.5 +/- 5.4-fold (P less than 0.005) above that in untreated rats. In the same experiment E2 alone had no significant effect on hepatic IGF-I gene expression. Similarly, in chronic experiments uterine IGF-I in ovx-hypox rats receiving 0.1 or 1 microgram/rat.day E2 for 10 days was significantly increased compared to that in ovx-hypox rats that did not receive E2 [5.38 +/- 0.79 vs. 1.10 +/- 0.15 (P less than 0.005) and 6.64 +/- 0.28 vs. 0.93 +/- 0.06 (P less than 0.005), respectively]. While administration of human GH alone significantly increased uterine IGF-I expression (3.76 +/- 1.61-fold compared to that in untreated rats; P less than 0.05), a significant and reproducible attenuation of E2-induced IGF-I expression was seen in the two acute experiments where GH reduced the E2-induced response by 36 +/- 3.7% and 53 +/- 19.4%. While chronic administration of E2 to ovx-hypox rats resulted in uterine growth, a significant decrease in body weight was seen in rats treated with 1 microgram/day E2 compared to that in untreated ovx-hypox controls (-4.3 +/- 1.5 vs. 2.5 +/- 0.6 g; P less than 0.0005). E2 treatment also significantly decreased the GH-induced increase in weight gain at each GH dose by approximately 40%. GH-induced hepatic IGF-I gene expression and serum IGF-I concentration were similarly reduced by chronic E2 administration. In contrast, in acute experiments where E2 alone had no effect on hepatic IGF-I expression, it acted in a synergistic fashion with GH and resulted in significantly greater accumulation of IGF-I mRNA. From these observations we conclude that 1) both E2 and human GH are potent stimulators of IGF-I gene expression in appropriate target tissues; and 2) in addition to any effects E2 has on GH secretion and IGF-I action, the growth-retarding effect of estrogen in the rat involves inhibition of GH-dependent hepatic IGF-I expression.

Animals↗

PubMed1988

Stimulation of hepatic insulin-like growth factor-I gene expression by ovine prolactin: evidence for intrinsic somatogenic activity in the rat.

The effect of ovine PRL (oPRL) on hepatic insulin-like growth factor I (IGF-I) gene expression was examined in the hypophysectomized (hypox) rat and compared with that seen in hypox rats treated with ovine GH (oGH) and human GH (hGH). A single injection of oPRL (100 micrograms/100 g BW) resulted in a 15-fold increase in hepatic IGF-I messenger RNA abundance 12 h after administration. Serum IGF-I increased from 82.9 +/- (SE) 9.0 to 137.0 +/- 15.4 ng/ml. In acute studies, where the slopes of 4-point dose-response curves were compared, oPRL was approximately 50% as potent as oGH in stimulating serum IGF-I and hepatic IGF-I messenger RNA accumulation. In chronic experiments, where a single daily injection of oPRL (0, 10, 30, or 90 micrograms/day) was administered for 10 days, a significant increase in body wt was observed at each dose. However, in contrast to hypox rats which received oGH, no dose-response relationship was apparent, and oPRL was less than 10% as potent as oGH in this regard. A small but significant increase in serum IGF-I and hepatic IGF-I expression was observed in hypox rats chronically injected with oPRL when killed 6-7 h after the final injection. However, by extrapolation from the hGH dose-response curves, oPRL was 6.8% and 22.7% as potent as hGH in stimulating serum IGF-I and hepatic IGF-I accumulation respectively. The observations reported here demonstrate that oPRL has significant somatogenic activity in the rat as determined by the three variables measured. Whereas oPRL had significant effects on body weight gain, serum IGF-I and hepatic IGF-I expression, the potency of oPRL by comparison with the two other somatogenic hormones was more marked in acute experiments compared to the effects seen after chronic administration. These results suggest that oPRL, like human GH, has both somatogenic and lactogenic activity in the rat and therefore may be an inappropriate control in studies of the effects of somatogenic hormones in the rat.

Animals↗

PubMed1988

Immunological studies on teleocalcin and salmon corpuscles of Stannius.

This report describes the characterization of an antiserum to salmon teleocalcin, a glycoprotein hormone that has recently been isolated from sockeye salmon, Oncorhynchus nerka, corpuscles of Stannius (CS). Immunodiffusion studies showed that teleocalcin was immunologically identical in four species of Pacific salmon. Histological staining of adjacent sections of sockeye salmon CS by periodic acid-Schiff reagent and immunocytochemistry revealed that the teleocalcin-immunoreactive cells were also periodic acid-Schiff positive. In bioassays using rainbow trout (Salmo gairdneri), the biological effects of teleocalcin were completely abolished by pretreatment with the antiserum before injection. The antiserum was ineffective by itself, however, in neutralizing endogenous teleocalcin. Immunoprecipitation studies of in vitro translation products from salmon CS poly(A)+ RNA identified a 26K protein, which corresponds to the size of the teleocalcin monomer as predicted by cDNA sequencing studies. Similar banding patterns were obtained with purified teleocalcin and crude CS extracts by Western blot analysis. However, there was one extra band (32K) in the CS extracts that was not present in the purified hormone preparations. This may represent the pro-form of the hormone in salmon.

Animals↗

PubMed1988

Role of calcium in prolactin-stimulated c-myc gene expression and mitogenesis in Nb2 lymphoma cells.

Receptor-activated transmembrane calcium flux has been implicated as a mediator of the actions of many growth factors and hormones. We examined the effects of PRL, calcium ionophores, and calcium antagonists on 45Ca2+ flux, c-myc gene expression, and DNA synthesis in the PRL-dependent rat Nb2 lymphoma cell line. PRL had no detectable effects on 45Ca2+ uptake or efflux, and the mitogenic effects of PRL could not be reproduced by the calcium ionophore A23187 alone or in combination with the tumor-promoting phorbol ester 12-O-tetra-decanoyl-phorbol-13 acetate (TPA). PRL, but not A23187 or TPA, stimulated c-myc gene expression in quiescent Nb2 cells. Exposure to PRL for brief periods (15 min to 4 h), followed by extensive washing, resulted in a time- and dose-dependent activation of DNA synthesis measured 16 h later. This activation was not blocked by addition of excess anti-PRL antiserum after the wash steps, indicating that the observed stimulation was not due to residual PRL. Despite the marked increase in DNA synthesis, removal of PRL after 4 h prevented mitosis, suggesting that PRL may be required throughout the cell cycle for Nb2 cell proliferation. Although continuous incubation with calcium antagonists resulted in a dose-dependent inhibition of PRL-stimulated DNA synthesis, activation of DNA synthesis by brief exposure to PRL was not inhibited by the presence of EGTA, calcium channel blockers (nifedipine, cobalt chloride), or calmodulin inhibitors (trifluoperazine, N-6-aminohexyl-5-chloronaphthalene sulfonamide). PRL-stimulated c-myc expression was attenuated, but not blocked, by the calcium channel antagonists. However, the putative intracellular calcium antagonist TMB-8 inhibited both c-myc expression and DNA synthesis in a dose-dependent manner (IC50 = 16 microM). Nb2 cells were sensitive to TMB-8 throughout G1 of the cell cycle, but inhibition of DNA synthesis was greatest when TMB-8 was present during the first 3 h of mitogen presentation, indicating a block in the transition from G0 to G1 of the cell cycle. The effect of TMB-8 suggests that release of a small intracellular calcium pool may mediate the early actions of PRL in Nb2 cells.

Animals↗

PubMed1988

A human B-lymphoblastoid cell line produces prolactin.

A variety of cell lines were examined by Northern blot hybridization for the expression of PRL or PRL-related mRNAs. We found that a human B-lymphoblast cell line transcribed a mRNA which hybridized to human PRL cDNA under high stringency conditions. The human lymphoblast cell line of interest is a variant subline of the IM-9 line that we have designated IM-9-P. The lymphoblast-derived PRL mRNA is approximately 150 bases longer than that produced by the human pituitary as determined by Northern blot analysis. IM-9-P PRL was immunoaffinity purified from conditioned medium and found to be identical in mol wt by sodium dodecyl sulfate-polyacrylamide gel electrophoresis to human pituitary PRL. Moreover, IM-9-P PRL is biologically active in the rat Nb2 lymphoma mitogenic assay. Ribonuclease-H digestion of mRNA poly(A) tracts indicated that the size difference between pituitary and IM-9-P PRL transcripts was not due to an elongated poly(A) tail on the lymphoid PRL mRNA. Genomic Southern blot analysis showed no major rearrangements of the PRL gene in IM-9-P cells compared to the parent IM-9 line and human placenta DNA. Thus, it is highly likely that an elongation of the 5' and/or 3' untranslated regions of IM-9-P PRL mRNA account for the size difference with pituitary PRL mRNA. The PRL-producing IM-9-P line was cloned by limiting dilution, and a high PRL-producing clone IM-9-P3 and a non-PRL producer IM-9-P6 were isolated for further analysis. IM-9-P3 cells were found to secrete 40-50 ng PRL/10(6) cells.24 h regardless of cell density. The level of PRL mRNA also remained constant during exponential growth of IM-9-P3 cells. The existence of the PRL-producing IM-9-P3 clone and the IM-9-P6 clone which does not produce PRL as well as the IM-9 progenitor line provides a unique system with which to analyze the molecular mechanism of ectopic human PRL expression.

Antibodies, Monoclonal↗

PubMed1988

Development of a primary culture system for rainbow trout corpuscles of stannius and characterization of secreted teleocalcin.

A primary culture system has been established for rainbow trout corpuscles of Stannius (CS). A RIA has also been developed to monitor and characterize the teleocalcin secreted by these cultures. The primary cultured CS cells actively synthesized and secreted teleocalcin for up to 39 days when maintained in amphibian and mammalian culture media. Numerous teleocalcin cells were also evident after immunocytochemical staining of the CS cultures. When the cultures were labeled with L-[35S]methionine, de novo synthesized and secreted teleocalcin was judged to be a glycosylated protein on the basis of Concanavalin-A-Sepharose binding and was similar in size to the intracellular form of the hormone. This culture system may prove to be ideal for identifying those factors that regulate teleocalcin secretion.

Animals↗

PubMed1988

Glycosylation selectively alters the biological activity of prolactin.

We have undertaken studies to determine the effect of glycosylation on the lactogenic activity of ovine PRL (oPRL). Measuring casein production in the in vitro mouse mammary gland explant assay, we found that glycosylated oPRL had 80% of the activity of oPRL. In competitive binding studies using lactogen receptors from mammary glands of lactating rabbits, glycosylated oPRL had only 20% the potency of oPRL. In the Nb2 assay also, glycosylated oPRL was approximately 24% as potent as oPRL in stimulating mitogenic activity. Thus, these studies show that the glycosylated variant of PRL has less biological activity than the major PRL form and that the alteration of an activity by glycosylation is selective.

Animals↗

PubMed1988

A third prolactin-like protein expressed by the developing rat placenta: complementary deoxyribonucleic acid sequence and partial structure of the gene.

We have previously reported on the isolation and characterization of two PRL-related cDNA clones, rat placental lactogen II and rat PRL-like protein A which are among the more abundant mRNAs expressed int he late term rat placenta. In this paper we report the isolation and characterization of cDNA clones of a third abundant placental protein, the predicted amino acid sequence of which shows a 44% homology to rPRL. This protein which we have called rPRL-like protein B (rPLP-B) is different from, but related to, the other PRL related proteins which have been identified in the rat, mouse, and bovine placentas. Two rPLP-B mRNA transcripts of 0.9 and 1.2 kilobases are strongly expressed in essentially equal amounts from day 14 of pregnancy until term. Nucleotide sequence and hybrid select translation data predict one secreted, potentially glycosylated protein of approximately mol wt 27,000. Hybridization and primer extension studies show that the two transcripts differ in their 5'-untranslated regions. One of the cDNA clones isolated represents a portion of the unprocessed rPLP-B mRNA. All intron/exon boundaries in this clone are of the same splice class and occur in identical locations within the coding region as in the rPRL gene.

Amino Acid Sequence↗

PubMed1988

Growth hormone-releasing hormone stimulates and somatostatin inhibits the release of a novel protein by cultured rat pituitary cells.

We have reported that the secretion of at least 17 distinct peptides [including rat (rGH)] GH by cultured rat pituitary cells was stimulated by GH-releasing hormone and inhibited by somatostatin, when analyzed by two-dimensional polyacrylamide gel electrophoresis. Three of these peptides (no. 23, 24, and 25) were not rGH immunoreactive. In order to determine whether these three peptides are fragments, degradation products or posttranscriptionally modified forms of rGH, rGH and peptide no. 23 were characterized structurally. From partial peptide maps of rGH and peptide no. 23 by V8 protease or chymotrypsin, it appeared that these peptides were not related to each other. By N-terminal microsequencing of two-dimensional polyacrylamide gel electrophoresis purified peptide, we have obtained the sequence of 24 N-terminal amino acid residues of peptide no. 23. This sequence has no significant homology with rGH or any other reported protein sequence. Antiserum was generated against a synthetic oligopeptide corresponding to amino acid residues 3-24 of peptide no. 23. The antiserum cross-reacted with peptides no. 23, 24, and 25 upon Western blot analysis. These results indicate that peptide no. 23 has a novel structure unrelated to other pituitary hormones. Since its secretion is influenced by GH-releasing hormone and somatostatin, peptide no. 23 may represent a previously unrecognized structurally unique growth factor.

Amino Acid Sequence↗