Stretches of poly(adenylic acid) are added post- transcriptionally to most eucaryotic messenger RNAs. This modification is thought to be involved in the regulation of stability and/or translation of mRNAs. Poly(A) chains are subject to both shortening and lengthening in the cytoplasm, but little is known about the mechanisms for, and significance of, changes in poly(A) length on messenger RNAs. This research is directed at understanding the role of poly(A) and its complex metabolism in the function of messenger RNA. The metabolism of the poly(A) tail on tubulin mRNAs can be altered by manipulating protein synthesis in the flagellated eucaryote, Chlamydomonas reinhardii. Newly developed methods for nuclear transformation of Chlamydomonas permit an analysis of mRNA structural features which affect poly(A) metabolism. The role of the 3' untranslated end (3'UT) of mRNA in conferring differential sensitivity to poly(A) loss is being investigated by the introduction of tubulin (and other) genes with altered 3'UT's into cells by transformation. The hypothesis that poly(A) loss is influenced by ribosome distribution on an mRNA is being tested by introducing upstream stop codons into tublin genes. The protein composition of the poly(A)-ribonucleoprotein complex on tubulin mRNAs is being characterized by crosslinking poly(A) chains and their associated proteins with ultraviolet light. It will be determined whether this structure is altered at times when poly(A) metabolism is altered. The role of altered poly(A) metabolism in the stability and translation of tubulin, and other, mRNAs will be explored, using both in vivo (transformation) and in vitro methods.%%% This project investigates how the living cell regulates the levels of important structural and "housekeeping" proteins. Protein synthesis is a complex and highly regulated cell activity about which much research is centered. Very little is known about how the cell controls the breakdown and repair of its essential proteins. This research begins to fill that gap.***//
