Regulation of Apoptosis by the Interaction of Casp-3 with PKC and Small Heat Shock Proteins

细胞凋亡或程序性细胞死亡在多细胞生物体的正常发育中起着至关重要的作用，并为细胞防御病原体和致癌因子提供了一种机制。在动物细胞中，细胞凋亡是由一组保守的半胱氨酸蛋白酶执行的，称为半胱氨酸酶。了解caspase活性开启和关闭的机制是当今生物学中的一个基本问题。蛋白激酶CD(PKCd)和小分子热休克蛋白27(Hsp27)与caspase-3相互作用。PKCd使caspase-3磷酸化，增强其活性，初步结果表明Hsp27抑制caspase-3活性。此外，在caspase-3激活过程中，Hsp27的亚细胞定位发生了变化，这种变化依赖于PKCd。本项目将研究三元caspase-3：PKCd：Hsp27和/或二聚体Casp-3：hasp27复合体在调节caspase-3活性中起关键作用的假设。这些实验将首先确定Hsp27抑制caspase-3活性的机制。研究将集中在探索Hsp27是否抑制导致caspase-3激活或其蛋白酶活性的蛋白降解过程。下一步，本项目将确定PKCd对Hsp27的磷酸化是否影响Hsp27的抑制活性。其次，这些研究将考察三元和二聚体复合体的形成，以及PKCd活性如何影响这些蛋白质在体内的分布和结合。总之，这些研究将提供关于翻译后修饰和蛋白质-蛋白质相互作用控制多细胞生物体中细胞生死所必需的过程的基本机制的关键信息。多斯夫博士培养了几名本科生，其中一些人是少数族裔。这些学生中的一些人在国际会议上展示了他们的研究成果，并与人合著了几本出版物。通过这个项目，多斯夫博士将能够让几名代表不足的少数族裔学生参与到研究的各个方面，并将继续参与大学吸引代表不足的少数族裔学生在科学领域的努力。

Apoptosis or programmed cell death plays an essential role in normal development and provides a mechanism of cellular defense against pathogens and oncogenic factors in multicellular organisms. In animal cells, apoptosis is performed by a group of conserved cysteine proteases, named caspases. Understanding the mechanisms by which caspase activity is turned on and off poses a fundamental question in biology today. Protein Kinase Cd (PKCd) and the small heat shock protein 27 (hsp27) interact with caspase-3. PKCd phosphorylates caspase-3 enhancing its activity and preliminary results suggested that hsp27 inhibits caspase-3 activity. Moreover, the subcellular localization of hsp27 changes during caspase-3 activation in a PKCd-dependent fashion. This project will study the hypothesis that the ternary caspase-3:PKCd:hsp27 and/or a dimeric casp-3:hasp27 complexes play a key role in the regulation of caspase-3 activity. These experiments will establish first the mechanism by which hsp27 inhibits caspase-3 activity. Studies will focus on exploring whether hsp27 inhibits the proteolytic processing leading to the activation of caspase-3 or its protease activity. Next, this project will determine whether the phosphorylation of hsp27 by PKCd affects the inhibitory activity of hsp27. Second, the studies will examine the formation of the ternary and dimeric complexes and how the PKCd activity affects the distribution and association of these proteins in vivo. Together, these studies will provide crucial information on the basic mechanisms by which posttranslational modifications and protein-protein interactions control a process essential to the life and death of cells in multicellular organisms. Dr. Doseff has trained several undergraduate students some of whom are underrepresented minorities. Some of these students have presented their research results in international meetings as well as co-authored several publications. Through this project, Dr. Doseff will be able to involve several underrepresented minority students in various aspects of the study and will continue to participate in the university's effort to attract underrepresented minority students in the sciences.