Mechanisms of neurotrophin signaling through the p75 receptor

描述（申请人提供）：在哺乳动物神经系统发育过程中，p75神经营养因子受体在调节神经元存活和死亡之间的微妙平衡中起关键作用。此外，它还与许多涉及异常凋亡的神经病理有关，如阿尔茨海默病、脊髓病变和兴奋性毒性损伤。像TNF受体超家族的其他成员一样，p75激活一个分叉信号，促进细胞存活和细胞死亡。这些途径是如何被激活和差异调节来决定细胞的最终命运的，目前还不清楚。该建议的总体目标是阐明p75神经营养因子受体介导细胞死亡的分子机制。我们之前已经证明TRAF6和NRIF是p75介导的细胞凋亡所必需的两个受体相互作用蛋白。TRAF6是一种E3泛素连接酶，已知可通过TNF和toll样受体超家族的许多成员调节NF-kB和JNK的激活。相反，NRIF是一种新的DMA结合蛋白，与p75的胞内结构域（ICD）结合，因此提出了这个相互作用物如何进入细胞核以及p75是否调节NRIF的转录活性的问题。我们的初步数据表明，NRIF被TRAF6泛素化，这促进了其核易位。此外，p75最近被证明可以通过γ分泌酶进行膜内蛋白水解，从而导致ICD的释放。这一过程如何影响信号传导尚不清楚；然而，我们的初步数据表明，这是受体诱导细胞凋亡所必需的。类似的裂解已被证明可以介导许多跨膜蛋白（如Notch和APR）的核信号传导。因此，我们假设p75的裂解释放NRIF，通过traf6介导的泛素化作用将NRIF靶向到细胞核，从而调控基因转录并导致细胞凋亡。为了验证这一假设，我们提出确定(1)p75切割与其信号转导之间的关系，(2)受体蛋白水解在NRIF核穿梭中的作用，(3)NRIF泛素化的功能后果，以及(4)NRIF转录活性响应p75激活的变化。鉴于p75所涉及的病理条件越来越多，我们必须开始了解这种受体如何介导细胞死亡，以便制定干预策略。

DESCRIPTION (provided by applicant): The p75 neurotrophin receptor plays a key role in regulating the delicate balance between neuronal survival and death during the development of the mammalian nervous system. In addition, it has been implicated in a number of neuropathologies involving abnormal apoptosis, such as Alzheimer's disease, spinal cord lesion and excitotoxic injury. Like other members of the TNF receptor superfamily, p75 activates a bifurcating signal that promotes both cell survival and cell death. How these pathways are activated and differentially regulated to determine the ultimate fate of a cell is not well understood. The overall objective of this proposal is to elucidate the molecular mechanisms by which the p75 neurotrophin receptor mediates cell death. We previously demonstrated that TRAF6 and NRIF are two receptor interacting proteins essential for p75- mediated apoptosis. TRAF6 is an E3 ubiquitin ligase known to regulate activation of NF-kB and JNK by many members of the TNF and Toll-like receptor superfamilies. In contrast, NRIF is a novel DMA binding protein that associates with the intracellular domain (ICD) of p75, thus raising the question as to how this interactor gets to the nucleus and whether p75 regulates NRIF's transcriptional activity. Our preliminary data indicate that NRIF is ubiquitinated by TRAF6 and this promotes its nuclear translocation. In addition, p75 was recently shown to undergo intramembrane proteolysis by gamma-secretase, leading to the release of the ICD. How this processing affects signaling is not clear; however, our preliminary data indicate that this is required for the receptor to induce apoptosis. A similar cleavage has been shown to mediate nuclear signaling for a number of transmembrane proteins (e.g. Notch and APR). Therefore, we hypothesize that p75 cleavage releases NRIF, which is targeted to the nucleus by TRAF6-mediated ubiquitination, resulting in the regulation of gene transcription and subsequently apoptosis. To test this hypothesis we propose to determine (1) the relationship between p75 cleavage and its signal transduction, (2) the role of receptor proteolysis in the nuclear shuttling of NRIF, (3) the functional consequence of NRIF ubiquitination, and (4) the change in NRIF's transcriptional activity in response to p75 activation. Given the ever-growing number of pathological conditions where p75 has been implicated, it is imperative that we begin to understand how this receptor mediates cell death in order to develop intervention strategies.