Cell integrity and morphogenesis in a budding yeast septin mutant

The non-sporulating diploid strain V327 of Saccharomyces cerevisiae was pre viously isolated in a search for thermosensitive autolytic mutants. This pr eviously isolated rn a search Tor thermosensitive autolytic mutants. Int st rain is very efficient at releasing intracellular proteins into the medium when incubated at high temperatures. The expression of this lytic phenotype depends on a morphogenetic defect, consisting of the appearance of elongat ed chains of cells. Transmission electron microscopy revealed a mislocaliza tion of septa at semi-permissive temperatures and a total lack of septation together with abnormal cell wail architecture at a non-permissive temperat ure. The septin-encoding CDC10 gene was cloned by complementation of the pl eiotropic phenotype of the V327 mutant. Rescue and sequencing of CDC10 alle les from V327 revealed a point mutation that created a single amino acid ch ange in a region which is well conserved among septins. This new allele was named cdc10-11. The construction of a cdc10-11 haploid strain by substitut ing the CDC10 gene with the rescued allele permitted further genetic analys es of the mutation and allowed the construction of new homozygous cdc10-11 diploid strains that showed a reduced ability to sporulate. Fusing both the wild-type and the cdc10-11 alleles to green fluorescent protein (GFP) demo nstrated that the mutation does not affect the localization of this septin to the bud neck at the standard growth temperature of 24 degrees C, althoug h the morphogenetic phenotype at 37 degrees C parallels the disappearance o f Cdc10-GFP at the ring encircling the septum area.