A functional analysis of the Candida albicans homolog of Saccharomyces cerevisiae VPS4.
To investigate the role of the prevacuolar secretion pathway in the trafficking of vacuolar proteins in Candida albicans, the C. albicans homolog of the Saccharomyces cerevisiae vacuolar protein sorting gene VPS4 was cloned and analyzed. Candida albicans VPS4 encodes a deduced AAA-type ATPase that is 75.6% similar to S. cerevisiae Vps4p, and plasmids bearing C. albicans VPS4 complemented the abnormal vacuolar morphology and carboxypeptidase missorting in S. cerevisiae vps4 null mutants. Candida albicans vps4Delta null mutants displayed a characteristic class E vacuolar morphology and multilamellar structures consistent with an aberrant prevacuolar compartment. The C. albicans vps4Delta mutant degraded more extracellular bovine serum albumin than did wild-type strains, which implied that this mutant secreted more extracellular protease activity. These phenotypes were complemented when a wild-type copy of VPS4 was reintroduced into its proper locus. Using a series of protease inhibitors, the origin of this extracellular protease activity was identified as a serine protease, and genetic analyses using a C. albicans vps4Deltaprc1Delta mutant identified this missorted vacuolar protease as carboxypeptidase Y. Unexpectedly, C. albicans Sap2p was not detected in culture supernatants of the vps4Delta mutants. These results indicate that C. albicans VPS4 is required for vacuolar biogenesis and proper sorting of vacuolar proteins.
Lee
Samuel A
SA
Division of Infectious Diseases, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, USA. lees7@utscsa.edu
Jones
Jason
J
Khalique
Zachary
Z
Kot
John
J
Alba
Mercedes
M
Bernardo
Stella
S
Seghal
Alfica
A
Wong
Brian
B
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016282:Aspartic Acid Endopeptidases / Q000378:metabolism; D002176:Candida albicans / Q000201:enzymology* / Q000235:genetics*; D002268:Carboxypeptidases / Q000378:metabolism; D003001:Cloning, Molecular; D017124:Conserved Sequence / Q000235:genetics; D056827:Endosomal Sorting Complexes Required for Transport; D005656:Fungal Proteins / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D005816:Genetic Complementation Test; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D056827:Endosomal Sorting Complexes Required for Transport; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; C107528:VPS4 protein, S cerevisiae; D002268:Carboxypeptidases; D016282:Aspartic Acid Endopeptidases; C090753:SAP2 protein, Candida; D000251:Adenosine Triphosphatases