LOCALIZATION OF THE SCHIZOSACCHAROMYCES-POMBE RHO1P GTPASE AND ITS INVOLVEMENT IN THE ORGANIZATION OF THE ACTIN CYTOSKELETON

The Schizosaccharomyces pombe rho1p GTPase directly activates the (1-3 )beta-D-glucan synthase and participates in the regulation of cell wal l growth and morphogenesis in this fission yeast, Indirect immunofluor escence experiments using rho1p tagged with hemagglutinin have reveale d that rho1p was located at the growing tips during interphase and at the septum prior to cytokinesis, localising to the same areas as actin patches. In S. pombe cdc10-129 mutant cells, arrested in G(1), HA-rho 1p accumulates at one tip whereas in cdc25-22 mutants, arrested in G(2 ), HA-rho1p accumulates at both tips. In tea1-1 and tea2-1 cdc11-119 m utant cells, HA-rho1p is localised to the new growing tips, Overexpres sion of different rho1 mutant alleles caused different effects on cort ical actin patch distribution, (1-3)beta-D-glucan synthase activation, and sensitivity to cell wall specific antifungal drugs. These results indicate that multiple cellular components are activated by rho1p. Ov erexpression of the dominant negative rho1T20N allele was lethal as wa s the rho1(+) deletion. Moreover, when rho1(+) expression was represse d in actively growing S. pombe, cells died in about 10 to 12 hours. Un der these conditions, normal cell morphology was maintained but the le vel of (1-3)beta-D-glucan synthase activity decreased and the actin pa tches disappeared, Most cells lysed after cytokinesis during the proce ss of separation, and lysis was not prevented by an osmotic stabiliser . We conclude that rho1p localisation is restricted to growth areas an d regulated during the cell cycle and that rho1p is involved in cell w all growth and actin cytoskeleton organisation in S. pombe.