Genetic and physical interactions in Arabidopsis between developmentally regulated GTPases, DRG-family binding proteins and ski-like RNA helicases
Marcec, Matthew Jerome
MetadataShow full item record
DRG proteins are encoded by an ancient highly conserved family of GTP binding protein genes. DRG1 and DRG2 orthologs have been found in all eukaryotic organisms tested. Recent evidence in yeast and Arabidopsis suggests that another gene SLH1 a DEAD-Box Helicase, genetically interacts with DRG1 and DRG2. Likewise, the DRG Family Regulatory Proteins DFRP1 and DFRP2 genetically and physically interact with their respective DRG partners. DFRP1 and DFRP2 also interact genetically with SLH1. A triple mutation of either drg1, drg2 and slh1 or dfrp1, dfrp2 and slh1 produces an embryonic lethal phenotype. This study has two aims. First, to characterize the several double and triple mutant combinations of the drg, dfrp, and slh1 and second, to construct a 6-His tagged SLH1 recombinant fusion protein so that antibodies could be made, which could be used, to further study the SLH1 proteins physical interaction with the DRG and DFRP proteins and with ribosomes.;In order to characterize phenotypic changes in several mutations involving the DRG, DFRP, and SLH1 genes were obtained. First, double mutants were generated. A second group of mutants was generated in which the plant is homozygous for 2 mutant alleles and segregating for the third (i.e. drg1, DRG2/drg2, slh1). Preliminary studies showed that such plants that such plants had a phenotype in which one quarter to one half of the seeds were aborted. To determine if this abortion occurs due to a deficiency in the gametophyte or sporophyte stage of the plants life cycle, crosses between these mutant plants and wild type plants were made. These experiments showed that the triple mutant pollen and embryo sacs have a reduced viability.;The SLH1 DEAD-Box Helicase consists of roughly 2100 amino acid residues. Rather than raising an antiserum against the entire protein, we chose to target two conserved regions. These antisera recognize the cloned proteins and are specific for the regions they were designed for. They will be of value in studying possible physical associations between SLH1, DRG and DFRP proteins.