Association of DRG1 and DRG2 with Ribosomes from Pea, Arabidopsis and Yeast

Abstract

DRGs are highly conserved GTP binding proteins.All eukaryotes examined contain DRG1 and DRG2 orthologs. The first experimental evidence for GTP binding by a plant DRG1 protein and by DRG2 from any organism is presented. DRG1 antibodies recognized a single ~43-kDa band in plant tissues, whereas DRG2 antibodies recognized ~45-,43-,and 30-kDa bands.An in vitro transcription and translation assay suggested that the 45-kDa band represents full-length DRG2 and that the smaller bands are specific proteolytic products. Homogenates from pea roots and root apices were used to produce fractions enriched in cytosolic and microsomal monosomes and polysomes. DRG1 and the 45- and 43-kDa DRG2 bands occurred in the cytosol and associated with cytosolic monosomes.I n contrast,the 30-kDa form of DRG2 was strongly enriched in polysome fractions.Thus,DRG1and the larger forms of DRG2 may be involved in translational initiation, and the 30-kDa form of DRG2 may be involved in translational elongation.DRG1 and the 45-and43-kDa forms of DRG2 can reassociate with ribosomes in vitro, a process that is partially inhibited by GTP-y-S Cells expressing FLAG-tagged ribosomal proteins from transgenic lines of Arabidopsis and yeast also demonstrated DRG-ribosome interactions.