Summary: The non-sporulating diploid strain V327 of Saccharomyces cerevisiae was previously isolated in a search for thermosensitive autolytic mutants. This strain 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 elongated chains of cells. Transmission electron microscopy revealed a mislocalization of septa at semi-permissive temperatures and a total lack of septation together with abnormal cell wall architecture at a non-permissive temperature. The septin-encoding CDC10 gene was cloned by complementation of the pleiotropic phenotype of the V327 mutant. Rescue and sequencing of CDC10 alleles from V327 revealed a point mutation that created a single amino acid change 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 substituting the CDC10 gene with the rescued allele permitted further genetic analyses 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) demonstrated that the mutation does not affect the localization of this septin to the bud neck at the standard growth temperature of 24 °C, although the morphogenetic phenotype at 37 °C parallels the disappearance of Cdc10-GFP at the ring encircling the septum area.
AdamsA.,
PringleJ.1984; Relationship of actin and tubulin distribution to bud growth in wild-type and morphogenetic mutant Saccharomyces cerevisiae.. J Cell Biol 98:934–945
AlvarezP.,
SampedroM.,
MolinaM.,
NombelaC.1994; A new system for the release of heterologous proteins from yeast based on mutant strains deficient in cell integrity.. J Biotechnol 38:81–88
BotsteinD.,
AmbergD.,
MulhollandJ.,
HuffakerT.,
AdamsA.,
DrubinD.,
StearnsT.1997; The yeast cytoskeleton.. In The Molecular and Cellular Biology of the Yeast Saccharomyces: Cell Cycle and Cell Biology pp. 1–90PringleJ.R.,
BroachJ.R.,
JonesE.W.
Edited by Cold Spring Harbor, NY:
CidV.J.,
DureánA.,
del ReyF.,
SnyderM.,
NombelaC.,
SánchezM.1995; Molecular basis of cell integrity and morphogenesis in Saccharomyces cerevisiae.. Microbiol Rev 59:345–386
CidV.J.,
CenamorR.,
SánchezM.,
NombelaC.1998; A mutation in the Rhol-GAP-encoding gene BEM2 of Saccharomyces cerevisiae affects morphogenesis and cell wall functionality.. Microbiology 144:25–36
De la FuenteJ.M.,
ÁlvarezA.,
NombelaC.,
SánchezM.1992; Flow cytometric analysis of Saccharomyces cerevisiae autolytic mutants and protoplasts.. Yeast 8:39–45
DeMariniD.J.,
AdamsA.E.M.,
FaresH.,
De VirgilioC.,
ValleG.,
ChuangJ.S.,
PringleJ.R.1997; A septin-based hierarchy of proteins required for localised deposition of chitin in theSaccharomyces cerevisiae cell wall.. J Cell Biol 139:75–93
De VirgilioC.,
DeMariniD.,
PringleJ.R.1996; SPR28, a sixth member of the septin gene family in Saccharomyces cerevisiae that is expressed specifically in sporulating cells.. Microbiology 142:2897–2905
DiDomenicoB.J.,
BrownN.H.,
LupisellaJ.,
GreeneJ.R.,
YankoM.,
KoltinY.1994; Homologs of the yeast neck filament associated genes: isolation and sequence analysis of Candida albicans CDC3 and CDC10.. Mol Gen Genet 242:689–698
FaresH.,
GoetschL.,
PringleJ.R.1996; Identification of a developmentally regulated septin and involvement of the septins in spore formation in Saccharomyces cerevisiae.. J Cell Biol 132:399–411
FordS.K.,
PringleJ.R.1991; Cellular morphogenesis in the Saccharomyces cerevisiae cell cycle: localisation of the cdc11 gene product and the timing of events at the budding site.. Dev Genet 12:281–292
GiotL.,
KonopkaJ.B.1997; Functional analysis of the interaction between Afrlp and the Cdcl2p septin, two proteins involved in pheromone-induced morphogenesis.. Mol Biol Cell 8:987–998
HaarerB.,
PringleJ.R.1987; Immunofluorescence localisation of the Saccharomyces cerevisiae CDC12 gene product to the vicinity of the lOnm filaments in the mother-bud neck.. Mol Cell Biol 7:3678–3687
KimH.B.,
HaarerB.K.,
PringleJ.R.1991; Cellular mor-phogenesis in the Saccharomyces cerevisiae cell cycle: localisation of the CDC3 gene product and the timing of events at the budding site.. J Cell Biol 112:535–544
KinoshitaM.,
KumarS.,
MizoguchiA.,
IdeC,
HaraguchiT.,
HiraokaY.,
NodaM.1997; Nedd5, a mammalian septin, is a novel cytoskeletal component interacting with actin-based structures.. Genes Dev 11:1535–1547
LiuH.,
KrizekJ.,
BretscherA.1992; Construction of a GAL1- regulated yeast cDNA expression library and its application to the identification of genes whose overexpression causes lethality in yeast.. Genetics 132:665–673
LongtineM.S.,
DeMariniD.,
ValencikD.,
Al-AwarO.S.,
FaresH.,
De VirgilioC.,
PringleJ.R.1996; The septins: roles in cytokinesis and other processes.. Curr Opin Cell Biol 8:106–119
McKieJ.M.,
SutherlandH.F.,
HarveyE.,
KimU.J.,
ScamblerP.J.1997; A human gene similar to Drosophila melanogaster peanut maps to the DiGeorge syndrome region of 22qll.. Hum Genet 101:6–12
NakatsuruS.,
SudoK.,
NakamuraY.1994; Molecular cloning of a novel human cDNA homologous to CDC10 in Saccharomyces cerevisiae.. Biochem Biophys Res Commun 202:82–87
NeufeldT.P.,
RubinG.M.1994; The Drosophila peanut gene is required for cytokinesis and encodes a protein similar to yeast putative bud neck filament proteins.. Cell 77:371–379
NiedenthalR.K.,
RilesL.,
JohnstonM.,
HegemannJ.H.1996; Green fluorescent protein as a marker for gene expression and subcellular localisation in budding yeast.. Yeast 12:773–786
OrleanP.1997; Biogenesis of yeast wall and surface components.. In The Molecular and Cellular Biology of the YeastSaccharomyces: Cell Cycle and Cell Biology pp. 229–362PringleJ.R.,
BroachJ.R.,
JonesE.W.
Edited by Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
OzsaracN.,
BhattacharyyaM.,
ClancyM.J.1995; The SPR3 gene encodes a sporulation-specific homologue of the yeast CDC3/10/11/12 family of bud neck microfilaments and is regulated by ABFI.. Gene 164:157–162
SandersS.L.,
HerskowitzI.1996; The Bud4p protein of yeast, required for axial budding, is localised to the mother/bud neck in a cell cycle-dependent manner.. J Cell Biol 134:413–427
SikorskiR.S.,
HieterP.1989; A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae.. Genetics 122:19–27
SlaterM.L.,
BowersB.,
CabibE.1985; Formation of septumlike structures at locations remote from the budding sites in cytokinesis-defective mutants of Saccharomyces cerevisiae.. J Bacteriol 162:763–767