Regulation of Cellular Responsiveness to BDNF

描述（由申请人提供）：神经营养因子，脑源性神经营养因子（BDNF），通过调节神经元存活和突触可塑性在脑发育和功能中起关键作用。BDNF信号传导的缺陷可能导致发育、神经退行性和精神疾病。对BDNF的细胞反应由酪氨酸受体激酶trkB介导，其大小由活性全长trkB和截短的trkB同种型的相对水平决定，其作为BDNF信号传导的显性负抑制剂。 在皮层神经元的初步研究表明，全长的trkB的表达，但不是截断的trkB，刺激膜去极化通过增加细胞内Ca 2+的水平。还发现Ca 2+对trkB基因（TRKB）的两个启动子产生相反的影响，表明启动子的使用调节trkB亚型的表达。这些观察结果导致了工作假设：TrkB同种型表达，因此，神经元对BDNF的反应，是由Ca2+依赖性TRKB转录调节。 将使用重组DNA技术研究Ca 2+差异调节TRKB启动子的机制，以鉴定TRKB中的Ca 2+依赖性调节元件以及与它们相互作用的转录因子。将通过测量具有启动子特异性5 '非翻译序列的全长和截短的trkB mRNA的去极化诱导表达来研究Ca 2+介导的启动子利用在调节全长和截短的trkB的差异表达中的作用。 为了将钙依赖的TRKB调节在一个更生理的背景下，我们将调查的能力，电诱发的钙离子瞬变，模拟在体内神经元的活动，差异激活TRKB启动子和差异驱动全长和截短的trkB亚型的表达模式。 本研究的长期目标是阐明Ca2+调节trkB亚型表达的机制，从而调节发育中，成人和患病大脑中神经元对BDNF的反应。

DESCRIPTION (provided by applicant): The neurotrophin, brain derived neurotrophic factor (BDNF), plays a critical role in brain development and function by regulating neuron survival and synaptic plasticity. Defects in BDNF signaling may lead to developmental, neurodegenerative and psychiatric disorders. Cellular responses to BDNF are mediated by the tyrosine receptor kinase, trkB, and their magnitude is determined by the relative levels of active, full-length trkB, and of truncated trkB isoforms, which act as dominant-negative inhibitors of BDNF signaling. Preliminary studies in cortical neurons revealed that expression of full-length trkB, but not that of truncated trkB, is stimulated by membrane depolarization via increased levels of intracellular Ca2+. Ca2+ was also found to exert opposing effects on the two promoters of the trkB gene (TRKB), suggesting that promoter usage regulates trkB isoform expression. These observations have led to the working hypothesis: TrkB isoform expression and consequently, neuronal responsiveness to BDNF, is regulated by Ca2+-dependent TRKB transcription. The mechanism by which Ca2+ differentially regulates the TRKB promoters will be studied using recombinant DNA technology to identify the Ca2+-dependent regulatory elements in TRKB and the transcription factors hat interact with them. The role of Ca2+-mediated promoter utilization in regulating the differential expression f full-length and truncated trkB will be studied by measuring the depolarization-induced expression f full-length and truncated trkB mRNA with promoter-specific 5'-untranslated sequences. In order to place Ca2+-dependent TRKB regulation in a more physiological context, we will investigate the ability of patterns of electrically evoked Ca2+ transients, which mimic in vivo neuronal activity, to differentially activate the TRKB promoters and differentially drive expression of full-length and truncated trkB isoforms. The long-term goal of this research is to elucidate the mechanisms by which Ca2+ regulates expression of trkB isoforms and, thus, modulates neuronal responsiveness to BDNF in the developing, adult and diseased brain.