Although the regulation of gonadotropin subunit gene expression and secretion by GnRH pulse frequency has been well documented in whole animals and cell culture systems, the molecular basis for this differential regulation is not understood. Our goal in this proposal is to uncover the molecular mechanisms underlying the pulse sensitivity of the gonadotrope cell. We hypothesize that the pulse sensitivity is conferred by the balance of transcriptional activators and co-repressors on the LHB and FSHB promoters. We also hypothesize that the levels of the activators and repressers are modulated by the frequency of pulses via distinct second messenger pathways. Initially, we will investigate the transcription factors and co-repressors that target the LHP and FSH|3 promoters. We have evidence that both activators (Egr-1, AP-1) and co-repressors (DAX-1, NAB-2, TGIF, SnoN) are induced at specific pulse frequencies. We will modulate the expression of each of these factors using lentiviral overexpression or siRNA knockdown to test their involvement in pulse decoding. Secondly, we will test the hypothesis that the GnRH-R activates Gs and cAMP signaling selectively at low pulse frequencies, but also activates Gq/11 and DAG/calcium and ERK signaling at high pulse frequencies. These studies will use a combination of standard biochemical measurements and state-of-the-art real-time measurements in live cells using newly developed FRET reporters. We will manipulate individual proteins in these signal cascades by lentiviral overexpression or siRNA depletion to test the effect on both signaling and gonadotropin expression and secretion. We furthermore hypothesize that the steroid hormone milieu will alter the expression of elements of these pathways and, consequently, the response of the gonadotrope to pulses of GnRH. Lastly, we will create genetic models of disrupted pulse sensing through genetic manipulation in the mouse. These genetic models will include cell-specific transgenic overexpression of selected co-repressors as well as gonadotrope-specific deletion of components of the Gs and Gq/11 pathways using cre-loxP technology.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD047400-04
Application #
7420898
Study Section
Molecular and Cellular Endocrinology Study Section (MCE)
Program Officer
Lamar, Charisee A
Project Start
2005-07-01
Project End
2010-04-30
Budget Start
2008-05-01
Budget End
2009-04-30
Support Year
4
Fiscal Year
2008
Total Cost
$260,925
Indirect Cost
Name
Veterans Medical Research Fdn/San Diego
Department
Type
DUNS #
933863508
City
San Diego
State
CA
Country
United States
Zip Code
92161
Fernandez, Marina O; Sharma, Shweta; Kim, Sun et al. (2017) Obese Neuronal PPAR? Knockout Mice Are Leptin Sensitive but Show Impaired Glucose Tolerance and Fertility. Endocrinology 158:121-133
Tang, Kechun; Pasqua, Teresa; Biswas, Angshuman et al. (2017) Muscle injury, impaired muscle function and insulin resistance in Chromogranin A-knockout mice. J Endocrinol 232:137-153
Cho, Chang Gun; Pak, Kwang; Webster, Nicholas et al. (2016) Both canonical and non-canonical NF-?B activation contribute to the proliferative response of the middle ear mucosa during bacterial infection. Innate Immun 22:626-634
Sen, Supriya; Langiewicz, Magda; Jumaa, Hassan et al. (2015) Deletion of serine/arginine-rich splicing factor 3 in hepatocytes predisposes to hepatocellular carcinoma in mice. Hepatology 61:171-83
Sen, Supriya; Jumaa, Hassan; Webster, Nicholas J G (2013) Splicing factor SRSF3 is crucial for hepatocyte differentiation and metabolic function. Nat Commun 4:1336
Sharma, Shweta; Morinaga, Hidetaka; Hwang, Vicky et al. (2013) Free fatty acids induce Lhb mRNA but suppress Fshb mRNA in pituitary L?T2 gonadotropes and diet-induced obesity reduces FSH levels in male mice and disrupts the proestrous LH/FSH surge in female mice. Endocrinology 154:2188-99
Lan, Debin; Lu, Min; Sharma, Shweta et al. (2011) Trans-resveratrol inhibits phosphorylation of Smad2/3 and represses FSH? gene expression by a SirT1-independent pathway in L?T2 gonadotrope cells. Reprod Toxicol 32:85-92
Godoy, Joseph; Nishimura, Marin; Webster, Nicholas J G (2011) Gonadotropin-releasing hormone induces miR-132 and miR-212 to regulate cellular morphology and migration in immortalized LbetaT2 pituitary gonadotrope cells. Mol Endocrinol 25:810-20
Sharma, Shweta; Sharma, Prem M; Mistry, Devendra S et al. (2011) PPARG regulates gonadotropin-releasing hormone signaling in LbetaT2 cells in vitro and pituitary gonadotroph function in vivo in mice. Biol Reprod 84:466-75
Mistry, Devendra S; Tsutsumi, Rie; Fernandez, Marina et al. (2011) Gonadotropin-releasing hormone pulse sensitivity of follicle-stimulating hormone-beta gene is mediated by differential expression of positive regulatory activator protein 1 factors and corepressors SKIL and TGIF1. Mol Endocrinol 25:1387-403

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