Dissection of the Functions of N-terminal Arginylation

描述（由申请人提供）： 由泛素系统调节的蛋白质降解控制许多调节蛋白质的浓度。 作为Ub系统的一种途径，N-末端规则途径的靶标包括带有不稳定N-末端残基的蛋白质。 N端作用途径的功能包括染色体分离的保真度。因此，预期该途径的扰动会影响哺乳动物细胞谱系中恶性表型出现的频率。在N-末端作用的去稳定残基中，N-末端Asp、Glu和Cys是独特的，因为它们在通过N-末端规则途径识别底物蛋白之前与Arg缀合。N-末端Asp、Glu和Cys的N-末端酰化由ATE 1编码的Arg-tRNA蛋白转移酶介导。Varshavsky实验室已经证明，缺乏N-末端乙酰化的小鼠ATE 1-/-品系，由于心脏发育和血管生成重塑的缺陷，在胚胎时死亡。此外，发现N-末端Cys的N-酰化需要其预先氧化，这表明N-末端作用的N-酰化分支可能起氧传感器的作用。我将在小鼠中进行分子遗传学研究，使用允许在特定细胞谱系中有条件消融N-末端乙酰化的技术，详细剖析N-末端规则途径的这一主要分支的心血管和其他功能。我还将鉴定R-转移酶的生理底物并分析ATE 1-/-细胞中的染色体稳定性。

DESCRIPTION (provided by applicant): Regulated protein degradation by the ubiquitin system controls the concentrations of many regulatory proteins. The targets of the N-end rule pathway, one pathway of the Ub system, include proteins bearing destabilizing N-terminal residues. The functions of the N-end role pathway include fidelity of chromosome segregation. Perturbations of this pathway are therefore expected to influence the frequency of emergence of malignant phenotypes in the mammalian cell lineages. Among destabilizing residues in the N-end role, N-terminal Asp, Glu, and Cys are unique in that they are conjugated to Arg before the recognition of a substrate protein by the N-end rule pathway. The arginylation of N-terminal Asp, Glu, and Cys is mediated by ATE1-encoded Arg-tRNA protein transferases. The Varshavsky lab has demonstrated that mouse ATE1 -/- strains, lacking N-terminal arginylation, die as embryos, owing to defects in heart development and angiogenic remodeling. In addition, arginylation of N-terminal Cys was discovered to require its prior oxidation, suggesting that the arginylation branch of the N-end role may function as an oxygen sensor. I will carry out molecular genetic studies in the mouse, using techniques that allow conditional ablation of N-terminal arginylation in specific cell lineages, to dissect, in detail, the cardiovascular and other functions of this major branch of the N-end rule pathway. I will also identify physiological substrates of R-transferases and analyze chromosome stability in ATE1 -/- ceils.