Regulation of mRNA Stability in Plastids - a Genetic and Biochemical Approach

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9404841 Gruissem Plastids in higher plants respond to the developmental program and environmental signals by adjusting the transcription rate of genes and the processing and stability of their mRNAs. The change in plastid mRNA stability is an important regulatory mechanism that is of functional consequence for translational efficiency. Most plastid mRNAs have an inverted repeat (IR) sequence in their 3' untranslated region (UTR) that is required for correct mRNA 3' end processing and the stability of the mature mRNA. Mutagenesis or deletion of the 3' IR sequence causes incorrect processing and/or rapid decay of the mRNA. Five nuclear-encoded proteins have been identified that interact with the 3' UTR of several plastid mRNAs in a structural and/or sequence-dependent manner. This project will take a combined biochemical and genetic approach to investigate the function of each protein and their possible interactions. Al proteins will be tested individually or in combinations for their role in mRNA 3' end processing and stability using reconstituted or partially fractionated in vitro systems. These studies will be complemented by parallel antisense-RNA strategies using Arabidopsis to confirm the in vivo function of the proteins. Previous experiments have already shown that the 28RNP is critical for chloroplast development in leaves. Similar antisene-RNA strategies will be used to determine the developmental role of the 24RNP which differs in its expression pattern from the 28RNP. The 100RNP will be tested for its potential interaction with the 24RNP and 28RNP to form a mRNA processing complex. Other experiments will investigate the function of the 29RNP and 55RNP and the interaction of these proteins with the 3'UTR of the petD mRNA. Together the biochemical and genetic dissection of the RNA-binding proteins will provide novel information on the nuclear control of organelle gene expression. Because of the importance of photosynthesis for plant productivity, the basic research proposed in this application is likely to have relevance for future plant and agricultural technologies. %%% Development of chloroplasts and photosynthetic competence are critical for plant growth and function. It has been shown that plastids respond to the plant developmental program and environmental signals by adjusting the expression of genes encoded in the plastid genome. One important control of plastid gene expression during development occurs at the level of mRNA processing and stability. Changes in mRNA stability can directly affect the translation of proteins that have essential functions in photosynthesis and other chloroplast processes. Processing and stability of chloroplast mRNAs are mediated by nuclear-encoded proteins that interact with specific sequences or structures in the RNA molecule. In previous studies, five proteins have been identified that participate in the processing and stability of a specific class of chloroplast mRNAs. The current project will take a combined biochemical and genetic approach to investigate the function of each protein and their possible interactions. Together, the biochemical and genetic dissection of the mechanisms by which chloroplast mRNA stability is accomplished will provide novel information on the nuclear control of organelle gene expression. Because of the importance of photosynthesis for plant productivity, the basic research proposed in this project is likely to have relevance for future plant and agricultural technologies. ***