Cells transcribe mRNAs (messenger RNAs) from genes and make the proteins they need from information contained in the mRNA molecules. The amount of a particular mRNA in a cell depends on the relative rates of its synthesis and degradation at that time. The rate of production of the protein often depends upon the abundance of its mRNA - - derangements in mRNA levels can be severely deleterious to the health and development of the organism. Features of a mRNA that make it susceptible to degradation as well as features that stabilize it are to be identified. The array of mRNAs in a cell changes during normal development. The nature and the control of the cellular machinery for degrading and stabilizing an mRNA are to be investigated also. We will study the plastid gene rbcL of Chlamydomonas reinhardtii: A section of rbcL encodes a target for mRNA degrading activity; a different segment of the gene encodes a mRNA stabilizing element. The fundamental questions about cellular RNA metabolism to be asked are: What specific sequence (or sequences) in the destabilizing region provide a primary target for the degrading enzyme (or enzymes)? What is the functionally important sequence in the stabilizing region and how does it act to prolong the lifetime of the transcript? C. reinhardtii cells can be grown in light or in darkness. An interesting feature of rbcL mRNA metabolism is that the same transcript has a much shorter lifetime in illuminated than in unilluminated cells. For example, the half-life of a chimeric gene containing certain rbcL sequences is 15 times shorter in cells transferred into the light than growing in darkness. How the light signal is perceived and propagated to make such a difference is another subject of this project. Modified reporter genes will be introduced into the plastid chromosome of C. reinhardtii and the stabilizing and destabilizing regions of the mRNA will be defined in studies of these transformants. The other cellular machinery for controlling the stability of the transcripts will be investigated through the generation and analysis of nuclear gene mutants and through certain biochemical approaches.
