Hsp27 Association with Contractile Proteins

描述(申请人摘要)：本项目的目标是 了解HSP27在正常生理机制中的作用 结肠的平滑肌肉收缩。调查员提出了存在 结肠平滑肌中的细胞内信号级联导致 热休克蛋白27的磷酸化及其与其他蛋白的相互作用 收缩蛋白，特别是肌动蛋白、原肌球蛋白、肌球蛋白和钙调蛋白， 导致平滑的肌肉收缩。这个级联是由PKC发起的 通过PKC转位到膜上的激活，导致MAP的激活 激活剂。该假说认为HSP27在维持 细胞骨架的完整性，与细胞骨架收缩之间的联系 平滑肌细胞内的蛋白质。申请者的学习将决定如何 HSP27的磷酸化使其与收缩蛋白相关联， 导致收缩。新出现的数据表明，HSP27的磷酸化 在细胞骨架重组中发挥作用，通过增加稳定性和 平滑肌细胞中收缩蛋白的关联性。HSP27是 对蛙皮素等不同的收缩激动剂有反应的磷酸化， 内皮素、卡巴胆碱和神经酰胺。初步数据还表明， HSP27的磷酸化伴随着HSP27的络合增加 HSP27与肌动蛋白和原肌球蛋白结合。收缩还伴随着 HSP27与肌动蛋白和原肌球蛋白的强共定位。松弛药 神经肽VIP和PKC抑制剂Calphostin C都能抑制 收缩和抑制HSP27的磷酸化，表明 HSP27的磷酸化可能与收缩有关。的具体目标 本研究的目的是：1)研究PKC介导的收缩对心肌细胞的影响。 Hsp27磷酸化与细胞骨架重组：2)检测 PKC介导的收缩对HSP27与肌动蛋白结合的影响 收缩蛋白是Smooth中收缩机械的成员 肌肉，即肌球蛋白、钙调蛋白和原肌球蛋白；以及3)检查 HSP27的表达变化与肌动蛋白、肌球蛋白、 转基因细胞和转基因小鼠体内的钙调蛋白和原肌球蛋白。

DESCRIPTION (Applicant's Abstract): The objective of this project is to understand the role of HSP27 in normal physiologic mechanisms that modulate smooth muscle contraction of the colon. The investigator propose the existence of an intracellular signaling cascade in colonic smooth muscle that results in the phosphorylation of HSP27 and the association of HSP27 with other contractile proteins, particularly actin, tropomyosin, myosin and caldesmon, leading to smooth muscle contraction. This cascade is initiated by PKC activation, by PKC translocation to the membrane, leading to activation of MAP kinases. The hypothesis is that HSP27 plays an important role in maintaining cytoskeletal integrity, and cytoskeletal association between the contractile proteins within smooth muscle cells. The applicant's studies will determine how phosphorylation of HSP27 confers association with the contractile proteins, leading to contraction. Emerging data suggest that phosphorylation of HSP27 plays a role in cytoskeletal reorganization by increasing stability and association of the contractile proteins in smooth muscle cells. HSP27 is phosphorylated in response to different contractile agonists like bombesin, endothelin, carbachol and ceramide. Preliminary data also suggest that phosphorylation of HSP27 is accompanied by an increase in the complexing of HSP27 with actin and with tropomyosin. Contraction is also accompanied with a strong colocalization of HSP27 with actin and with tropomyosin. The relaxant neuropeptide VIP and the PKC inhibitor calphostin C cause both an inhibition of contraction and an inhibition of HSP27 phosphorylation, suggesting that phosphorylation of HSP27 and contraction may be related. The specific aims of this proposal are to: 1) examine the effect of PKC mediated contraction on HSP27 phosphorylation and cytoskeletal reorganization: 2) examine the effect of PKC mediated contraction on the association of HSP27 with actin, and other contractile proteins that are members of the contractile machinery in smooth muscle, namely myosin, caldesmon, and tropomyosin; and 3) examine the effect of altered expression of HSP27 on the association of HSP27 with actin, myosin, caldesmon and tropomyosin in transfected cells and in transgenic mice.