An important characteristic of cellular senescence is the loss of proliferative capacity. During this terminal mitotic phase, fibroblasts exhibit characteristic morphological changes with accompanying alterations in gene expression. Gene expression in eukaryotic cells is regulated to a significant extent by post-transcriptional modification of pre-mRNA, which includes polyadenylation, cleavage and alternative splicing. Although post-transcriptional processing has been investigated in other systems, its relevance in cellular aging has not been examined. We have identified alterations in the levels and activities of proteins which bind to heterogeneous nuclear RNA (hnRNA) in senescent fibroblasts; these are the hnRNP A1 and A2 proteins and both are functional in the biogenesis and stability of mature mRNA. The altered activities are expected to produce several effects for specific RNAs, such as reduced mRNA, defective proteins, and inefficient translation. All are potential mechanisms to induce cellular senescence. Our hypothesis is that post-transcriptional processing has an important role in affecting gene expression during cellular senescence. The goals of this proposal are to investigate the age-related roles of hnRNP A1 and A2 proteins in modulating gene expression. We will assess changes in post-transcriptional processing in senescent cells by using RNA binding and splicing assays. We expect to find that lifespan will be altered by manipulating the expression of hnRNP A1 and A2 proteins. We will individually overexpress hnRNP A1 and A2 proteins in normal human fibroblasts to determine their effects on lifespan using an inducible retroviral system. Based on our preliminary findings, we anticipate that lifespan will be lengthen. The effects on lifespan would be the result of hnRNP A1 and A2 modulating the expression of specific RNA species. We will identify these RNAs by the use of differential display. Another important activity of hnRNP A1 and A2 is the binding to telomere sequences. This activity has only been demonstrated in vitro, therefore, we will also determine if telomere length is affected in cells overexpressing A1 and A2 proteins. The long term goal of the studies will be to further characterize the genes identified by differential display and investigate the mechanisms by which A1 and A2 regulate their expression. The studies proposed will assess the contribution post-transcriptional processing has in modulating gene expression during cellular aging.
NIH ExPorter
NIH ExPorter