9303898 Peltz Recently, it has become clear that the decay rates of mRNAs are regulated and play an important role in controlling gene expression. Work from many laboratories has demonstrated the requirement for translation in the degradation of mRNAs, and understanding the relationship between these two processes is a major challenge for this field. One clear example of this relationship is the observation that nonsense mutations accelerate the decay rates of mRNAs 10- to 20-fold. We have demonstrated that the product of the UPF1 gene is involved in this pathway. Deletion of the UPF1 gene from the yeast chromosome completely abrogates the accelerated decay of nonsense-containing mRNAs without affecting the decay of wild-type transcripts. The DNA sequence of the UPF1 gene has been determined and demonstrated to contain several sequence motifs and regions of homology with other genes that may suggest several putative activities, including: 1) an NTP binding and hydrolysis motif(s), 2) an RNA helicase motif, 3) Zinc-finger motifs and 4) sequence homology with the SEN1 gene, whose protein product is the noncatalytic subunit of the tRNA splicing endonuclease. Biochemical analysis of the Upf1 protein has revealed that it is polysome-associated and that the decay rates of nonsense-containing mRNAs in extracts prepared from UPF1+ and upf1- strains recapitulate the results observed in cells. Based on these observations, the objective of this proposal is to genetically characterize the UPF1 gene in order to begin to identify its role in the nonsense-mediated mRNA decay pathway. Mutations in the UPF1 gene will be isolated and their alleles will be characterized to determine the effect of these mutations on the decay of nonsense- containing mRNAs. %%% Proper cellular growth and differentiation is determined by the regulated expression of the genetic material within the cell. The control of gene expression has been studied intensively in many organisms and ha s been demonstrated to be controlled at multiple points from transcriptional initiation to protein stability. Recently, it has become clear that the decay rates of mRNAs are regulated and play an important role in controlling gene expression. The decay rates of mRNAs vary from each other by more than fifty-fold. In addition, mRNA decay can be modulated depending upon multiple factors including the stage of the cell cycle, cellular differentiation and reponses to hormones. The mechanisms which are responsible for determining either the intrinsic decay rate of an mRNA or the regulation of that rate are largely unknown. It is the goal of my laboratory to identify both cis-acting sequences and trans-acting factors which regulate mRNA stability. ***
