Regulation of the MAP Kinase Pathway in Senescence

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DESCRIPTION (provided by applicant): Replicative senescence is characterized by a general dysregulation of numerous cellular processes. However, the hallmark of senescent cells in vitro is failure to respond to mitogens by DNA synthesis and cell division. Our previous studies have shown that failure of the proliferative response is based primarily in signaling failures and these occur downstream of receptor binding. The MAP Kinase pathway is the pathway most associated with the mitogenic response. We have examined the abundance and activity of various intermediates in this pathway. Although there were some changes in mitogen stimulated increases in ERK activity between young and senescent cells, when calculated per mg of protein, the activities were the same. The most dramatic change in senescence was the much reduced abundance of p-ERK in the nucleus of senescent cells. This was consistent with the failure of senescent cells to phosphorylate p-ERK?s target, Elk-1. The experiments proposed in this project will address the mechanism of failed translocation of nuclear p-ERK. We will examine whether dimerization of p-ERK required for transport, is defective in senescence. We will examine phosphatase activity in the nucleus of young and senescent cells. We will determine whether there is accelerated export of p-ERK from the nucleus and whether there is failure of a nuclear anchor(s) in senescent cells. In addition, ?we will determine the consequences of restoring p-ERK in senescent cells and finally we will use SV40 T antigen as a probe of the mechanisms underlying the macromolecular assembly of nuclear p-ERK. Our goal is to understand the mechanisms underlying the failure in MAP Kinase mitogen signal transduction in culture as a model for signaling failures in aging.