Poster: Protein targeting & vesicular trafficking
Abs #
651: Functional Studies of the Tic Machinery in Arabidopsis
Chloroplast biogenesis is dependent on the efficient import of nuclear-encoded proteins from the cytosol. Precursor proteins carrying an N-terminal transit peptide are post-translationally translocated across the outer- and inner-envelope membranes by hetero-oligomeric protein complexes termed Toc and Tic, respectively. Most components of the Toc and Tic were initially identified in vitro as pea chloroplast envelope proteins found in close proximity to translocating precursors. Here we show that Arabidopsis homologues of two Tic components, Tic40 and Tic110, and of a stromal chaperone, Hsp93, all play a role in chloroplast protein import in vivo. Arabidopsis knockout mutants for the unique atTic40 and atTic110 genes, as well as for one of the two genes encoding chloroplast Hsp93, atHsp93-V, were found to display clear chlorotic phenotypes and chloroplast ultrastructural defects. These were further shown to be associated with defects in chloroplast protein import, providing strong evidence for a role of these proteins in the import process. Protein import efficiency was evaluated in these mutants using the traditional in vitro import approach as well as novel in vivo assays. Because Tic40 has previously been shown to have significant levels of homology with the Hip and Hop co-chaperones, different atTic40 deletion and fusion constructs were tested for complementation of tic40 in order to evaluate the functional significance of this homology. In addition, various genetic and biochemical approaches have been taken to provide supporting evidence for a functional cooperation between these three components in a putative stromal import motor. Results from the Arabidopsis mutant characterization, tic40 complementation and functional cooperation studies will be presented.
