POLY (A) FNCTN & METABOLISM IN SACCHAROMYCES CEREVISIAE

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How genes are differentially regulated by altering the metabolism of their mature mRNA is a key question in molecular biology. Potential points of regulation include nuclear-cytoplasmic mRNA transport, mRNA stability, and mRNA translational efficiency. The poly(A) tail of mRNA is considered important for each of these processes, yet no clear understanding of poly (A) tail function exists. The long term objective of this work is to analyze the role(s) of poly(A) in mRNA metabolism and to understand how modifications in it or the protein bound to it, the poly(A)-binding protein, can affect this metabolism. With this understanding, potential alterations of poly(A)-dependent steps in mRNa metabolism resulting from cellular infection or transformation can be studied. The initial objective of the work is to characterize genes in Saccharomyces cerevisiae which affect poly(A) synthesis, packaging, and degradation. These include genes encoding a poly(A)-polymerase, the poly(A)-binding protein, and seven genes (spbl-spb7) which when mutated allow the cells to live without the essential poly(A)-binding protein. One of these, SPB2, encodes the ribosomal protein L46, and another, SPB4, encodes a protein which is highly homologous to eucaryotic initiation factor 4a and is probably an RNA-helicase. The effects of mutations in these genes on in vivo mRNA transport, poly(A) and mRNA stability, and mRNA translation will be examined. An in vitro poly(A)-degradation assay that faithfully reproduces the in vivo effects of these mutations will be developed. This assay will detect the functional interactions between the poly(A) tail and the remainder of the mRNA, and it will be a model system for studying messenger ribonucleoprotein structure and its affects on mRNA stability.