BASIS FOR STRESS TOLERANCE IN OSTEOLIGAMENT CELLS

描述：(改编自申请人的摘要)假设 本申请所基于的翻译是- α1(I)前胶原链的易位是由一种 转位蛋白(内质多跨膜糖蛋白 和内质网(ER)驻留的分子伴侣。 此外，前胶原与个体易位的关系 蛋白质和一系列分子伴侣遵循 连续的动作。根据申请人的学习情况，候选人 对于这种与前胶原的连续相互作用，被认为可能 包括转运子相关蛋白(TRAP)，转移链- 结合膜蛋白(TRAM)是SEC61p的哺乳动物同源物， Hsp47(一种特异性胶原结合蛋白)、GRP78和Grp94。 转位蛋白被认为起到调节结合的作用。 核糖体参与合成前胶原，其靶向是 ER膜由信号识别颗粒及其受体组成。 随着新生链条的转移，它们被保护起来不受 膜的疏水核心由转位组分组成。这个 新生的前胶原链然后被递给一组连续的 分子伴侣，Hsp47，GRP78和Grp94，促进 通过减少错误折叠来减少翻译-移位过程，从而 确保翻译-移位高保真地发生。这 在韧带细胞中，蛋白质-蛋白质相互作用的级联作用确保 前胶原在正常和正常状态下的成分合成 压力。这一假设将通过完成以下工作来检验 明确的目标。特定目标1将验证转运子包括 转运链结合膜蛋白与SEC61p的关系 存在于韧带细胞和成纤维细胞的内质网中 前胶原I.特异性目标2将鉴定哺乳动物的成分 结合翻译α1的核糖体的内质网(I) 前胶原mRNA，并与α(I)转位有关 新生的前胶原链。具体的重点将指向有轨电车， SEC61p和HSP47。《特定目标3》将试图证明 单个转位蛋白之间的连续关联，ER 常驻分子伴侣和前胶原蛋白。利用交联剂 和重建方法将确定HSP47是否是 第一个与进化中的内质网驻留分子伴侣相互作用 前胶原链并确定前胶原合成点 与GRP78、Grp94和其他内质网驻留蛋白相关。

DESCRIPTION: (Adapted from the applicant's abstract) The hypothesis upon which this application is based is that the translation- translocation of alpha1(I) procollagen chains is facilitated by a translocon (multispaning membrane glycoproteins of the endoplasmic reticulum), and endoplasmic reticulum (ER) resident molecular chaperons. Further, the association of procollagen with individual translocon proteins and a series of molecular chaperons follows the principle of successive action. Based on the studies of the applicant, candidates for this consecutive interaction with procollagen are seen to possibly include the translocon-associated protein (TRAP), translocating chain- associating membrane protein (TRAM), the mammalian homolog of SEC61p, Hsp47 (a specific collagen binding protein), Grp78, and Grp94. Translocon proteins are proposed to act to mediate the binding of ribosomes engaged in synthesizing procollagen that has been targeted to the ER membrane by the signal recognition particle and its receptor. As translocation of the nascent chains proceeds, they are shielded from the hydrophobic core of the membrane by translocon components. The nascent chains of procollagen are then handed to a successive group of molecular chaperons, Hsp47, Grp78, and Grp94, that facilitate the translation-translocation process by reducing incorrect folding, thus insuring that translation-translocation occur with high fidelity. This cascade of protein-protein interactions ensures, in ligament cells, the constitutive synthesis of procollagen during periods of normalcy and stress. This hypothesis will be tested by completion of the following specific aims. Specific Aim 1 will verify that a translocon comprised of translocating chain associating membrane protein (TRAM) and SEC61p exists in the ER of ligament cells and fibroblasts that produce procollagen I. Specific Aim 2 will identify components of the mammalian endoplasmic reticulum that bind ribosomes which translate alpha1(I) procollagen mRNA, and are associated with translocation of alpha(I) procollagen nascent chains. Specific focus will be directed to TRAM, SEC61p and HSP47. Specific Aim 3 will attempt to prove that there is a successive association between individual translocon proteins, ER resident molecular chaperons and procollagen. Utilizing crosslinking and reconstitution methods it will be determined whether Hsp47 is the first ER resident molecular chaperone to interact with evolving procollagen chains and determine the point of procollagen synthesis that is associated with Grp78, Grp94 and other ER resident proteins.