MAIZE RIBOSOMAL PROTEIN GENE STRUCTURE AND EXPRESSION

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The long-term objectives of the proposed research are to understand how gene structure relates to gene expression; how the cell regulates the expression of large sets of genes to produce functional plastid (70S) and cytosolic (80S) ribosomes; and how the expression of plastid- and nuclear-encoded 70S ribosomal protein genes is coordinated in the separate genomes. A large increase in the cellular content of 70S and 80S ribosomes occurs during maize leaf cell development. The specific aims are to thoroughly characterize the ribosome content throughout maize leaves; determine the content of 80S and plastid-encoded and nuclear- encoded 70S ribosomal protein mRNAs in leaves using ribosomal protein cDNAs as hybridization probes; and to examine the rates of ribosomal protein synthesis in the leaves. Additionally, the influence of 70S ribosome content on the expression nuclear- encoded 70S ribosomal protein gene expression will be studied in mutants of maize which pre cold-sensitive for the accumlation of the 70S ribosomes. These experiments are designed to determine the mechanisms leading to the balanced accumulation of 70S ribosomal proteins. Complementary to the above studies will be the investigation of 80S and nuclear-encoded 70S ribosomal protein gene structure. cDNAs for these genes be isolated using heterologous 70S and 80S ribosomal protein cDNAs. The maize cDNAs will then be used in the above experiments and to select the genomic clones from a maize library. Several 80S and nuclear-encoded 70S ribosomal protein genes will be sequenced. Transcription start and stop sites will be identified by S1 nuclease mapping and primer extension analysis. Computer-assisted sequence analysis of these genes may reveal DNA sequences that could be involved in the coordinate regulation of the ribosomal protein genes. Protein binding activity of the genes will be examined by DNA footprint analysis. Additionally, the possibility that some ribosomal proteins bind to pre-mRNAs or mature mRNAs will be tested by protein-RNA binding assays.