Isolation, Characterization, and Regulation of the Prolactin Receptor

Vonderhaar, Barbara K.; Bhattacharya, Alok; Alhadi, Taufiek; Liscia, Daniel S.; Andrew, Elizabeth M.; Young, Janet K.; Ginsburg, Erika; Bhattacharjee, Maitreyi; Horn, Toby M.

doi:10.3168/jds.s0022-0302(85)80847-x

Cited by 19 publications

(17 citation statements)

References 136 publications

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“…A molecular weight of approximately 36,000 and 87,000 for the binding domain is consistent with affinity cross-linking studies using animal tissues [7][8][9]651 and SDS-PAGE analysis of affinity purified material identified by Western blot analysis [66-68 J or probing with 1251-hCH [62,67]. Unidentified bands of these molecular weights have also been seen on coomassie or silver stained gels of affinity purified material including human chorion [5,69].…”

Section: Discussionsupporting

confidence: 65%

Solubilization and characterization of a lactogenic receptor from human placental chorion membranes

Ormandy

Sutherland

1990

J of Cellular Biochemistry

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Prolactin has a wide range of actions, including osmoregulation and the control of mammary gland development and lactation. These effects are mediated through a high-affinity cell surface receptor, which has been well characterized in a number of animal tissues. The molecular characteristics of the human receptor are unknown, however. The present studies were initiated, therefore, to determine the binding and molecular characteristics of the lactogenic receptor of human placental chorion membranes. Subcellular fractionation studies showed that the bulk of the receptor sedimented in the microsomal fraction at 45,000gav. Endogenous ligand was dissociated from the receptor with 3.5 M MgCl2 or 0.05 M acetate buffer (pH 4.8) with preservation of binding activity. The microsomal receptor bound human growth hormone (hGH), human prolactin (hPRL), ovine prolactin (oPRL), and human placental lactogen (hPL) but not non-primate growth hormones, indicating a narrow specificity for lactogenic hormones. The binding was only partially reversible in agreement with the known binding kinetics of animal lactogenic receptors. The receptor was solubilized with 45% yield from the microsomes using 16 mM 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulphonate (CHAPS) detergent-250 mM NaCl, and the binding activity was fully restored by a two-fold dilution in the binding reaction to reveal a KD of 0.8 nM for hGH and a binding capacity of 200 fmol of specifically bound hGH per mg of microsomal protein. Gel filtration chromatography indicated the minimum molecular weight of the ligand-receptor complex was approximately 60,000 daltons, and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of covalently cross-linked 125I-hGH-receptor complexes revealed a molecular size of 58,000 daltons. When account was taken of the contribution of the ligand, a molecular weight of 36,000 for the receptor's binding domain was obtained. These data indicate that the chorion lactogenic receptor has very similar binding and molecular characteristics to the lactogenic receptors from other mammalian species. Chorion membranes are thus a convenient source of material for the further purification and characterization of the human lactogenic receptor.

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Section: Discussionsupporting

confidence: 65%

Solubilization and characterization of a lactogenic receptor from human placental chorion membranes

Ormandy

Sutherland

1990

J of Cellular Biochemistry

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“…We demonstrated that, during the increased secretion of hCG after 'A and 1 h, the percent age of radiolabeled hCG did not change significantly, either in the intracellular or in the extracellular compart ment. The decreased extracellular secretion of hCG be tween 2 and 24 h, however, was accompanied by a signifi cant decrease in radiolabeled hCG in both compartments at 2, 4, 6, 12 and 24 h. The existence of prolactin receptors in the human pla centa is known [11,12], The action of human prolactin which we observed in our experiments is probably recep tor mediated and therefore very specific. Ovine prolactin failed to have any effect on secretion of hCG.…”

Section: Influence O F Human Prolactin On De Novo Synthesis O F Hcgsupporting

confidence: 42%

Effects of Prolactin on Secretion and Synthesis of Human Chorionic Gonadotropin in Human Term Placentas in vitro: Short-Term Increase in Secretion, Followed by Medium-Term Suppression of Synthesis and Secretion

Würfel

Beckmann

Austin

et al. 1992

Gynecol Obstet Invest

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We studied the influence of human prolactin on the secretion and de novo synthesis of human chorionic gonadotropin (hCG) in the human term placenta in culture. Placental tissue from 14 patients with uncomplicated pregnancies and deliveries was prepared mechanically, with addition of a Percoll gradient step. hCG levels were determined in the culture media and in the cytosolic fraction of cells by means of an enzyme immunoassay with coated beads. The amount of newly synthesized hCG was measured by the extent of incorporation of ³⁵S-methionine into the hCG molecule. Our results showed that human prolactin had two different effects in vitro: between ½ and 1 h, prolactin slightly increased secretion of hCG into the culture medium without affecting de novo synthesis; after 2 h, prolactin began to cause a significant decrease in both secretion and de novo synthesis of hCG over several hours. It appears that both effects are receptor mediated, for ovine prolactin failed to produce any response. We conclude that prolactin is one of the main factors regulating the synthesis and secretion of hCG in the human trophoblast at term.

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“…Proteins were subjected to 10% SDS-PAGE, transferred to Hybond nitrocellulose ECL membrane and probed with mPrlR antibody (M2-5 for EpH4 cells; 1:250 dilution (Vonderhaar et al, 1985); or hPRLR antibody (Invitrogen; 2 μg/ml). Tubulin was used as loading control (Sigma).…”

Section: Western Blot Analysismentioning

confidence: 99%

Progesterone induces expression of the prolactin receptor gene through cooperative action of Sp1 and C/EBP

Goldhar

Duan

Ginsburg

et al. 2011

Molecular and Cellular Endocrinology

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Prolactin (Prl) and progesterone (P) cooperate synergistically during mammary gland development and tumorigenesis. We hypothesized that one mechanism for these effects may be through mutual induction of receptors (R). EpH4 mouse mammary epithelial cells stably transfected with PR-A express elevated levels of PrlR mRNA and protein compared to control EpH4 cells that lack the PR. Likewise, T47D human breast cancer cells treated with P overexpress the PrlR and activate PrlR promoter III. PrlR promoter III does not contain a classical P response element but contains several binding sites for transcription proteins, including C/EBP, Sp1 and AP1, which may also interact with the PR. Using promoter deletion and site directed mutagenesis analyses as well as gel shift assays, cooperative activation of the C/EBP and adjacent Sp1A, but not the Sp1B or AP1, sites by P is shown to confer P responsiveness leading to increased PrlR transcription.

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Isolation, Characterization, and Regulation of the Prolactin Receptor

Cited by 19 publications

References 136 publications

Solubilization and characterization of a lactogenic receptor from human placental chorion membranes

Solubilization and characterization of a lactogenic receptor from human placental chorion membranes

Effects of Prolactin on Secretion and Synthesis of Human Chorionic Gonadotropin in Human Term Placentas in vitro: Short-Term Increase in Secretion, Followed by Medium-Term Suppression of Synthesis and Secretion

Progesterone induces expression of the prolactin receptor gene through cooperative action of Sp1 and C/EBP

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