CREB and ICER, the Yin and Yang of FSH-beta regulation by pulsatile GnRH

描述（由申请人提供）：在我的博士后研究期间，我继续对生殖的神经内分泌控制和促性腺激素解码下丘脑GnRH神经元释放的脉动GnRH信号的机制感兴趣。这个奖项将提供机会，以满足我的长期目标，成为一个独立的调查员，补充领导，研究，并由布里格姆妇女医院提供的道德培训计划。垂体FSH和LH的产生，在脉冲式GnRH的控制下，对生育力至关重要。与LRH相反，FSH β亚基（FSH β）转录和FSH分泌优先在缓慢而非快速的脉动GnRH频率下刺激。这种差异的机制尚不清楚。本项目的目标是阐明促性腺激素释放激素脉冲频率依赖性差异控制FSH β转录和随后的FSH合成的潜在机制。使用我们的灌流系统和小鼠促性腺激素衍生的L β T2细胞系模型，我们先前已经表明，GnRH激活CREB介导在较慢的GnRH脉冲频率下优先诱导FSH β，而ICER在高GnRH脉冲频率下诱导并拮抗CREB的刺激作用。脉动GnRH调节CREB和ICER表达、激活和调节的分子机制仍然知之甚少。我建议延长我们以前在体外观察细胞系在体内小鼠模型，并预测促性腺激素特异性CREB和ICER缺陷的小鼠模型将进一步证明这些转录因子的关键功能，从下丘脑的GnRH信号解码，通过展示中断的周期性和受损的生殖功能。具体目标是确定两种转录因子，cAMP反应元件结合蛋白（CREB）和诱导型cAMP早期阻遏物（ICER），介导促性腺激素释放激素（FSH）对促性腺激素释放激素（GnRH）脉冲的差异反应，以促进生殖功能的方式。为了实现这些目标，我们提出了以下目标：1）研究GnRH脉冲频率对PKA活性和CREB磷酸化状态的调节机制，从而在慢频率和快频率下优先刺激FSH β转录; 2）研究GnRH脉冲频率依赖性地调节ICER合成、作用和降解的途径; 3）研究CREB和ICER基因缺失对促性腺激素细胞FSH β转录和生殖功能的影响。阐明促性腺激素释放激素（GnRH）调控FSH β转录的机制，可为多囊卵巢综合征（PCOS）和下丘脑性闭经（HA）等生殖系统疾病提供新的治疗靶点。

DESCRIPTION (provided by applicant): During my post-doctoral fellowship, I have continued my interest in the neuroendocrine control of reproduction and the mechanism by which the gonadotrope decodes the pulsatile GnRH signal, released from GnRH neurons in the hypothalamus. This award will provide the opportunity to meet my long-term goal of becoming an independent investigator, supplementing the leadership, research, and ethical training programs offered by Brigham and Women's Hospital. Production of pituitary FSH and LH, under the control of pulsatile GnRH, is essential for fertility. Pituitary FSHß-subunit (FSHß) transcription and hence FSH secretion are preferentially stimulated at slow rather than fast frequencies of pulsatile GnRH, in contrast to LHß. The mechanisms for this disparity are not known. The goal of this project is to elucidate the underlying mechanisms that contribute to the GnRH pulse frequency-dependent differential control of FSHß transcription and subsequent FSH synthesis. Using our perifusion system and the murine gonadotrope-derived LßT2 cell line model, we have shown previously that GnRH activation of CREB mediates preferential induction of FSHß at slower GnRH pulse frequencies, whereas ICER is induced at high GnRH pulse frequencies and antagonizes the stimulatory effects of CREB. The molecular mechanisms by which pulsatile GnRH regulates CREB and ICER expression, activation, and regulation remain poorly understood. I propose to extend our prior in vitro observations in cell lines to in vivo mouse models, and predict that gonadotrope-specific CREB and ICER deficient mouse models will further demonstrate the critical function of these transcription factors in decoding oscillatoy GnRH signals from the hypothalamus, by demonstrating disrupted cyclicity and impaired reproductive function. The specific objectives are to determine the manner in which two transcription factors, cAMP response element binding protein (CREB) and inducible cAMP early repressor (ICER), mediate the differential response of FSHß to pulsatile GnRH in the gonadotrope to contribute to reproductive function. To achieve these objectives, the following aims are proposed: 1) Investigate the mechanisms by which PKA activity and CREB phosphorylation status are regulated by pulsatile GnRH to preferentially stimulate FSHß transcription at slow versus fast frequencies; 2) Investigate the pathways that regulate ICER synthesis, action and degradation in a GnRH pulse frequency-dependent manner; 3) Study the effect of CREB and ICER gene disruption in gonadotropes on FSHß transcription and reproductive function in vivo. The elucidation of the mechanisms by which FSHß transcription is regulated by pulsatile GnRH could provide novel therapeutic targets for reproductive disorders such as polycystic ovarian syndrome (PCOS) and hypothalamic amenorrhea (HA).