A new genetic method for isolating functionally interacting genes: High plo1(+)-dependent mutants and their suppressors define genes in mitotic and septation pathways in fission yeast

We describe a general genetic method to identify genes encoding proteins th at functionally interact with and/or are good candidates for downstream tar gets of a particular gene product. The screen identifies mutants whose grow th depends on high levels of expression of that gene. We apply this to the ploI(+) gene that encodes a fission yeast homologic of the ploI(+)-dependen t (pld) mutants that show defects in mitosis or septation. Threee mutants s how a mitotic arrest phenotype. Among the 14 pld mutants with septation def ects, 12 mapped to known loci: cdc7, cdc15, cdc 11 spg1, and sid2. One of t he pld mutants cdc7-PDI, was seklected for suppressor analysis. As multicop y suppressors, we isolated foru known genes involved in septation in fissio n yeats: spg1(+), sce3(+), cdc8(+), and rhoI(+), and two previously unchara cterized genes, mpd1+ and mpd2(+) , mpd1(+) exhibits high homology to phosp hatidylinositol 4-phosphate 5-kinase, while mpd2(+) resembles Saccaromyces cerevisiae SMY2; both proteins are involved in the regulation of actin-medi ated processes. As chromosomal suppressors of cdc7-PD1, we isolated mutatio ns of cdc16 that resulted in multiseptation without nuclear division. cdc16 (+), dma1(+), byr4(+) and a truncated form of the cdc7 gene were isolated b y complementation of one of those cdc16 mutations. These results demonstrat e that screening for high dose-dependent mutants and their suppressors is a n effective approach to identify functionally interacting genes.