Chlamydomonas reinhardtii as a useful model system for the study of target of rapamycin (TOR) signalling in photosynthetic eukaryotes

Sandra Diaz-Troya, M. Esther Perez-Perez, Francisco J. Florencio and Jose L. Crespo

Instituto de Bioquimica Vegetal y Fotosintesis. Consejo Superior de Investigaciones Cientificas-Universidad de Sevilla, 41092 Seville, Spain

The highly conserved TOR kinase is a central controller of cell growth in response to nutrient availability in all eukaryotes. TOR exists in two functionally and structurally distinct complexes, termed TOR complex 1 (TORC1) and TORC2. TORC1, but not TORC2, is specifically inhibited by the complex formed by the immunosuppressive and anticancer drug rapamycin and the FKBP12 immunophilin. We identified homologues in Chlamydomonas to TOR, FKBP12, and the TORC1- and TORC2-shared protein LST8. Our data indicate that a rapamycin-sensitive signalling pathway composed by at least CrTOR, CrLST8 and CrFKBP12 participate in the regulation of Chlamydomonas cell growth. Biochemical fractionation and indirect immunofluorescence microscopy studies indicate that CrTOR and CrLST8 exist in high molecular mass complexes that associate with microsomal membranes and are particularly abundant in the peri-basal body region. A Saccharomyces cerevisiae complementation assay demonstrates that CrLST8 is able to functionally and structurally replace endogenous yeast LST8 and allows us to propose that binding of LST8 to TOR is essential for cell growth. We have addressed a proteomic and phosphoproteomic analysis of rapamycin-treated cells to find out the molecular mechanisms by which CrTOR controls Chlamydomonas cell growth. Based on our studies, we propose Chlamydomonas as a useful model system for investigating TOR signalling in photosynthetic eukaryotes because, unlike plants, Chlamydomonas is naturally sensitive to rapamycin.

e-mail address of corresponding author: eperez@ibvf.csic.es