PROTRH GENE TRANSCRIPTION AND BIOSYNTHESES BY LEPTIN

描述(申请人提供)：瘦素主要在脂肪组织中产生，在向调节食欲、能量消耗和神经内分泌功能的大脑中心提供有关能量储存和能量平衡的信息方面具有重要的生理作用。瘦素的作用之一是通过促甲状腺激素释放激素(TRH)肽激活下丘脑-垂体-甲状腺(HPT)轴，从而增加能量消耗。在过去三年的资助中，我们能够提供强有力的证据，证明瘦素在调节TRH前体激素生物合成方面存在直接(室旁核，PVN)和间接(弓状核，ARC)途径。我们证明瘦素可以直接调节培养的下丘脑神经元的前TRH的生物合成和TRH的分泌，而不依赖于黑素皮质素(α-黑素细胞刺激素，α-MSH)的输入。我们还观察到瘦素受体(ObRb)和前TRH在PVN神经元中的共存，强烈提示瘦素对TRH神经元的直接作用。进一步支持这一假说的是，在下丘脑室旁核中发现了ObRb的mRNA表达，并且在给大鼠注射瘦素后，TRH神经元表达了SOCS-3mRNA(这是瘦素直接作用的敏感标志)。最近，我们首次证明了瘦素在体内激活了TRH神经元中的STATS，进一步证明了STAT3转录因子在瘦素调节PRTRH启动子活性方面发挥了潜在的关键作用。我们最近还证明了瘦素作用引起的TRH生物合成的增加与参与TRH和其他PRTRH多肽成熟的加工酶PC2和PC2的上调有关。 目的#1我们将检验这样一种假设，即存在两组不同或重叠的PVN-TRH神经元，携带ObRb(直接通路)和MC4R(间接通路)，瘦素和α-MSH以此为靶点，通过增加HPT轴的能量消耗来调节能量平衡，并通过向LH和DMH发送突触来潜在地抑制食物的摄取。 目的#2我们将验证瘦素主要通过直接途径调节HPT轴的假设。 目的#3A)由于我们发现Leptin通过上调PC2和PC2的生物合成来调节proTRH并协调其加工，我们推测Leptin还可能调节其他酶、辅助伴侣和完全激素前体加工和成熟所需的抑制物。B)由于黑素皮质素系统是TRH神经元的间接作用途径，我们还将确定α-MSH是否影响PC2、ProSAAS、7B2和CPE的生物合成、成熟和活性。

DESCRIPTION (provided by applicant): Leptin, principally produced in adipose tissue, has a central physiologic role in providing information on energy stores and energy balance to brain centers that regulate appetite, energy expenditure and neuroendocrine function. One of the actions of leptin is to activate the Hypothalamic-Pituitary-Thyroid (HPT) axis through the thyrotropin-releasing hormone (TRH) peptide thereby increasing energy expenditure. During the last three years of funding, we were able to provide strong evidence for the existence of a direct (paraventricular nucleus, PVN) and an indirect (arcuate nucleus, ARC) pathway of leptin action on the regulation of the TRH prohormone biosynthesis. We demonstrated that leptin could directly regulate proTRH biosynthesis and TRH secretion in cultured hypothalamic neurons independently of the melanocortin (alpha-melanocyte-stimulating-hormone, alpha-MSH) input. We also observed the colocalization of the leptin receptor (ObRb) with proTRH in neurons of the PVN, strongly suggesting direct effects of leptin on TRH neurons. This hypothesis was further supported by the demonstration of ObRb mRNA expression within the PVN, and by showing that TRH neurons express SOCS-3 mRNA (which is the sensitive marker of direct leptin action) after leptin administration to rats. Recently, we demonstrated for the first time activation of STATS in TRH neurons by leptin in vivo, providing further evidence of a potential key role of the STAT3 transcription factor to regulate the activity of the proTRH promoter by leptin. We also recently demonstrated that the increase in TRH biosynthesis due to leptin action was associated with an up-regulation of the processing enzymes PCI and PC2, involved in the maturation of TRH and other proTRH peptides. Aim #1 We will test the hypothesis that there are two different or overlapping groups of PVN-TRH neurons carrying the ObRb (direct pathway) and the MC4R (indirect pathway) that are targeted by leptin and alpha-MSH to regulate the energy balance by increasing energy expenditure through the HPT axis, and potentially inhibiting food intake by sending synapses to the LH and DMH. Aim #2 We will test the hypothesis that leptin regulates the HPT axis primarily through the direct pathway. AIM #3 A) Because we found that leptin regulates proTRH and coordinates its processing by also up-regulating PCI and PC2 biosynthesis, we hypothesize that leptin might also regulate the other enzymes, co-chaperones, and inhibitors necessary for full prohormone processing and maturation. B) Since the melanocortin system represents the indirect pathway of action on TRH neurons, we will also determine whether alpha- MSH affects the biosynthesis, maturation and activity of PCI, PC2, proSAAS, 7B2 and CPE.