PHOSPHORYLATION MEDIATES THE NUCLEAR TARGETING OF THE MAIZE RAB17 PROTEIN

The maize abscisic acid-responsive Rab17 protein localizes to the nucl eus and cytoplasm in maize cells. In-frame fusion of Rab17 to the repo rter protein beta-glucuronidase (GUS) directed GUS to the nucleus and cytoplasm in transgenic Arabidopsis thaliana and in transiently transf ormed onion cells. Analysis of chimeric constructs identified one regi on between amino acid positions 66-96, which was necessary for targeti ng GUS to the nucleus. This region contains a serine cluster followed by a putative consensus site for protein kinase CK2 phosphorylation, a nd a stretch of basic amino acids resembling the simian virus 40 large T antigen-type nuclear localization signal (NLS), Mutation of two bas ic amino acids in the putative NLS had a weak effect on nuclear target ing in the onion cell system and did not modify the percentage of nucl ear fusion protein in the Arabidopsis cells. The mutation of three ami no acids in the consensus site for CK2 recognition resulted in the abs ence of in vitro phosphorylated forms of Rab17 and in a strong decreas e of GUS enzymatic activity in isolated nuclei of transgenic Arabidops is. These results suggest that phosphorylation of Rab17 by protein kin ase CK2 is the relevant step for its nuclear location, either by facil itating binding to specific proteins or as a direct part of the nuclea r targeting apparatus.