Effects of 8-bromo-cAMP on expression of endocrine functions by cultured human trophoblast cells. Regulation of specific mRNAs

Ringler, Guy E.; Kao, Lee-Chuan; Miller, Walter L.; Strauss, Jerome F.

doi:10.1016/0303-7207(89)90185-8

Cited by 85 publications

(22 citation statements)

References 23 publications

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“…In primary cultures, CG␣ and ␤ mRNA synthesis and secretion as well as transcription from the CG␣ promoter could be stimulated in response to cAMP (42,43,45). Addition of exogenous cAMP analogs was also shown to promote in vitro cell fusion of villous trophoblasts and resulted in down-regulation of the regulatory subunit of PKA type I (39,40).…”

Section: Discussionmentioning

confidence: 88%

“…During differentiation, an increase in CG secretion as a consequence of CG␣ and CG␤ mRNA de novo synthesis was observed reflecting the in vivo situation (41,42). CG␣ rose earlier than CG␤ during spontaneous differentiation or upon cAMP stimulation, suggesting that production of the latter is more strictly dependent on syncytium formation (43)(44)(45). We have recently demonstrated that the promoter of the CG␣ gene is active in isolated cytotrophoblasts and increases during in vitro differentiation (42).…”

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confidence: 99%

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Promoter Elements and Transcription Factors Involved in Differentiation-Dependent Human Chorionic Gonadotrophin- Messenger Ribonucleic Acid Expression of Term Villous Trophoblasts

Martin

2000

Endocrinology

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Differentiation of primary villous cytotrophoblasts into syncytia is associated with increasing production of ␣ and ␤ human CG subunits, which is predominantly governed at the level of messenger RNA expression. Here, we present a detailed study on the mechanisms involved in the differentiation-dependent regulation of the trophoblast-specific CG␣ gene promoter. Site-directed mutations in each of the five DNA-elements of the composite enhancer were performed to investigate the contribution of the individual regulatory sequences to the overall transcriptional activity of the promoter at two different stages of trophoblast in vitro differentiation. We show that deletion of one cyclic AMP response element (CRE) did not affect CG␣ promoter activity in cytotrophoblasts; however, it reduced transcription by 33% in differentiating cultures. Removal of both CREs almost abolished transcription at early and later stages of in vitro differentiation. Upon mutation the enhancer elements ␣ACT, JRE, and CCAAT significantly decreased luciferase reporter transcription; however their contribution to the total promoter activity did not change during in vitro differentiation. Contrary to that, mutated TSE diminished promoter activity by 19% during 12 and 48 h of cultivation but reduced luciferase expression by 78% between 48 and 84 h of differentiation. In electrophoretic mobility shift assay, the TSE interacted with activating protein (AP)-2␣ in both primary trophoblasts and choriocarcinoma cells. While CRE-interacting proteins were detectable 12 h after isolation, the TSE-binding complex did not appear before 36 h of in vitro differentiation. During syncytium formation increasing protein expression of activating transcription factor (ATF)-1, cAMP response element-binding protein (CREB)-1, and AP-2␣ was observed on Western blots. Moreover, phosphorylated CREB-1 and ATF-1 accumulated between 24 and 78 h of trophoblast cultivation. By fluorescence immunohistochemistry, we show that CREB-1 was predominantly expressed in syncytiotrophoblasts, whereas ATF-1 and AP-2␣ localized to the syncytium and some cytototrophoblasts as well as to stromal and endothelial cells of the placental villus. Phosphorylated CREB-1/ATF-1 and the coactivator protein CBP were primarily detected in syncytial nuclei, suggesting the presence of functional, cAMP-dependent transcriptional complexes in the differentiated tissue. In agreement to the in vivo situation, phosphorylated CREB-1/ATF-1 were observed in nuclei of the differentiated trophoblast cultures. The activity of the CG␣ promoter as well as CREB-1/ATF-1 phosphorylation increased upon elevation of cAMP levels and overexpression of the catalytic subunit of protein kinase A. Additionally, we demonstrate that overproduction of the enzyme enhanced protein expression and binding of AP-2␣ to the TSE. We conclude that differentiation-dependent transcription of the CG␣ gene in villous trophoblasts is mainly governed by increasing expression of AP-2␣ and PKA-dependent phosphorylation of CREB-1 and

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Section: Discussionmentioning

confidence: 88%

mentioning

confidence: 99%

Promoter Elements and Transcription Factors Involved in Differentiation-Dependent Human Chorionic Gonadotrophin- Messenger Ribonucleic Acid Expression of Term Villous Trophoblasts

Martin

2000

Endocrinology

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“…One group proposed that a short-lived, cycloheximide-sensitive DNA-binding protein, operationally termed steroid hormone inducing protein (SHIP) was involved in this process as a transcription factor (Waterman & Simpson 1989). While it soon became clear that the action of cycloheximide did not affect transcription of the steroidogenic enzyme genes or the accumulation of their encoded mRNAs (Golos et al 1987, Picado-Leonard et al 1988, Mellon & Vaisse 1989, Ringler et al 1989, it was still not clear exactly what cycloheximide was doing.…”

Section: Search For the Acute Regulator Of Steroidogenesismentioning

confidence: 99%

Congenital lipoid adrenal hyperplasia: the human gene knockout for the steroidogenic acute regulatory protein

Miller¹

1997

Journal of Molecular Endocrinology

132

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“…These observations suggest the in volvement of adenylate cyclase in regulation of basal and stimulated hCG secretion. cAMP stimulates hCG secretion from first-trimester [41,42] and term placental [43,44] and cho riocarcinoma cells [45], Similarly, cAMP in creased a-and (3-hCG mRNA transcription [46]. Here, GnRH and cAMP stimulated hCG secretion from first-trimester and term trophoblast cells ( fig.…”

Section: Hcg Secretory Response To Camp and Gnrh In Perifusionmentioning

confidence: 99%

Characterization of Cell Types within a Chorionic Gonadotropin-Secreting, Mechanically Dissociated Human Placental Cell Population in Perifusion

Steele

Currie²,

Yuen³

et al. 1993

Neurosignals

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A cell perifusion system was established to examine human placental endocrine regulation by locally synthesized peptides. First-trimester and term trophoblast cells were mechanically dissociated. Cells were plated on microcarrier beads and cultured for 7–14 days. Cells on beads were loaded in chambers, perifused with culture media and effluent was assayed for chorionic gonadotropin (hCG). Mechanical dissociation of placental tissue produced cell preparations with 85–95% viability. Staining with Masson trichrome, cytokeratin and β-hCG antibodies suggested that greater than 50% of the cells were trophoblast. Perifused trophoblast cells secreted hCG in a continuous non-pulsatile fashion, independent of exogenous hormonal stimuli. hCG secretion from first-trimester trophoblast cells remained stable in static culture for 14 days. GnRH perifusion (10^–8M) for 15–120 s transiently increased hCG secretion from first-trimester trophoblast cells. Longer GnRH exposure stimulated greater hCG secretion. Each of 3 consecutive pulses of 8-bromo-cyclic adenosine-3′,5′-monophosphate (cAMP, 10^–9M) administered at 2-hour intervals stimulated transient hCG secretion from first-trimester and term placental cells. cAMP stimulated hCG secretion more potently from first-trimester than from term placental cells.

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Effects of 8-bromo-cAMP on expression of endocrine functions by cultured human trophoblast cells. Regulation of specific mRNAs

Cited by 85 publications

References 23 publications

Promoter Elements and Transcription Factors Involved in Differentiation-Dependent Human Chorionic Gonadotrophin- Messenger Ribonucleic Acid Expression of Term Villous Trophoblasts

Promoter Elements and Transcription Factors Involved in Differentiation-Dependent Human Chorionic Gonadotrophin- Messenger Ribonucleic Acid Expression of Term Villous Trophoblasts

Congenital lipoid adrenal hyperplasia: the human gene knockout for the steroidogenic acute regulatory protein

Characterization of Cell Types within a Chorionic Gonadotropin-Secreting, Mechanically Dissociated Human Placental Cell Population in Perifusion

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