title: Candida albicans VMA3 is necessary for V-ATPase assembly and function and contributes to secretion and filamentation.
abstract: The vacuolar membrane ATPase (V-ATPase) is a protein complex that utilizes ATP hydrolysis to drive protons from the cytosol into the vacuolar lumen, acidifying the vacuole and modulating several key cellular response systems in Saccharomyces cerevisiae. To study the contribution of V-ATPase to the biology and virulence attributes of the opportunistic fungal pathogen Candida albicans, we created a conditional mutant in which VMA3 was placed under the control of a tetracycline-regulated promoter (tetR-VMA3 strain). Repression of VMA3 in the tetR-VMA3 strain prevents V-ATPase assembly at the vacuolar membrane and reduces concanamycin A-sensitive ATPase-specific activity and proton transport by more than 90%. Loss of C. albicans V-ATPase activity alkalinizes the vacuolar lumen and has pleiotropic effects, including pH-dependent growth, calcium sensitivity, and cold sensitivity. The tetR-VMA3 strain also displays abnormal vacuolar morphology, indicative of defective vacuolar membrane fission. The tetR-VMA3 strain has impaired aspartyl protease and lipase secretion, as well as attenuated virulence in an in vitro macrophage killing model. Repression of VMA3 suppresses filamentation, and V-ATPase-dependent filamentation defects are not rescued by overexpression of RIM8, MDS3, EFG1, CST20, or UME6, which encode positive regulators of filamentation. Specific chemical inhibition of Vma3p function also results in defective filamentation. These findings suggest either that V-ATPase functions downstream of these transcriptional regulators or that V-ATPase function during filamentation involves independent mechanisms and alternative signaling pathways. Taken together, these data indicate that V-ATPase activity is a fundamental requirement for several key virulence-associated traits in C. albicans.
authors.affiliation: Section of Infectious Diseases, New Mexico Veterans Healthcare System, Albuquerque, New Mexico, USA.
authors.forename: Hallie S
authors.initials: HS
authors.lastname: Rane
authors.forename: Stella M
authors.initials: SM
authors.lastname: Bernardo
authors.forename: Summer M
authors.initials: SM
authors.lastname: Raines
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Binder
authors.forename: Karlett J
authors.initials: KJ
authors.lastname: Parra
authors.forename: Samuel A
authors.initials: SA
authors.lastname: Lee
mesh_terms: D057055:Aspartic Acid Proteases / Q000378:metabolism; D002176:Candida albicans / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism / Q000472:pathogenicity; D005089:Exocytosis; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D008049:Lipase / Q000378:metabolism; D009154:Mutation; D055503:Protein Multimerization; D025262:Vacuolar Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*; D014617:Vacuoles / Q000378:metabolism / Q000648:ultrastructure; D014774:Virulence
chemical_list: D005656:Fungal Proteins; D008049:Lipase; D057055:Aspartic Acid Proteases; D025262:Vacuolar Proton-Translocating ATPases
references.citation: J Biol Chem. 2000 Dec 22;275(51):40088-95
references.pmid: 11007788
references.citation: Bioinformatics. 2005 Jun;21 Suppl 1:i251-7
references.pmid: 15961464
references.citation: J Gen Microbiol. 1987 Oct;133(10):2817-24
references.pmid: 3329678
references.citation: Yeast. 1997 Dec;13(15):1423-35
references.pmid: 9434348
references.citation: Commun Integr Biol. 2011 Mar;4(2):240-2
references.pmid: 21655451
references.citation: J Biol Chem. 2002 Oct 25;277(43):40544-8
references.pmid: 12186879
references.citation: Mol Cell Biol. 2010 Jul;30(14):3695-710
references.pmid: 20457806
references.citation: Eukaryot Cell. 2004 Dec;3(6):1609-18
references.pmid: 15590834
references.citation: Curr Protein Pept Sci. 2012 Mar;13(2):134-40
references.pmid: 22264127
references.citation: J Cell Biol. 1988 Oct;107(4):1369-83
references.pmid: 3049619
references.citation: Clin Infect Dis. 2004 Aug 1;39(3):309-17
references.pmid: 15306996
references.citation: J Bioenerg Biomembr. 2003 Aug;35(4):301-12
references.pmid: 14635776
references.citation: Cell Microbiol. 2005 Nov;7(11):1546-54
references.pmid: 16207242
references.citation: Mol Biol Cell. 2001 Nov;12(11):3631-43
references.pmid: 11694594
references.citation: Annu Rev Genet. 2003;37:435-60
references.pmid: 14616069
references.citation: Microbiol Rev. 1990 Sep;54(3):266-92
references.pmid: 2215422
references.citation: J Biol Chem. 2005 May 6;280(18):18393-402
references.pmid: 15718227
references.citation: J Biol Chem. 2013 Mar 1;288(9):6190-201
references.pmid: 23316054
references.citation: J Biol Chem. 2011 Mar 25;286(12):10058-65
references.pmid: 21239492
references.citation: J Biol Chem. 2006 Jan 6;281(1):90-8
references.pmid: 16263704
references.citation: J Biol Chem. 1999 Dec 24;274(52):37270-9
references.pmid: 10601292
references.citation: Yeast. 2011 Dec;28(12):833-41
references.pmid: 22072586
references.citation: Mol Microbiol. 2005 May;56(3):649-69
references.pmid: 15819622
references.citation: J Biol Chem. 2012 Mar 23;287(13):10236-10250
references.pmid: 22215674
references.citation: Nucleic Acids Res. 2005 Jul 1;33(Web Server issue):W289-94
references.pmid: 15980472
references.citation: J Exp Biol. 2002 May;205(Pt 9):1209-19
references.pmid: 11948198
references.citation: Mol Biol Cell. 2008 Apr;19(4):1354-65
references.pmid: 18216277
references.citation: FEMS Yeast Res. 2009 Feb;9(1):126-42
references.pmid: 19054126
references.citation: Microbiol Mol Biol Rev. 2003 Sep;67(3):400-28, table of contents
references.pmid: 12966142
references.citation: Proc Natl Acad Sci U S A. 1978 Jul;75(7):3327-31
references.pmid: 28524
references.citation: Eukaryot Cell. 2008 May;7(5):747-64
references.pmid: 18375617
references.citation: Trends Microbiol. 2001 Jul;9(7):327-35
references.pmid: 11435107
references.citation: J Biol Chem. 2011 Aug 12;286(32):28089-96
references.pmid: 21669878
references.citation: Eukaryot Cell. 2005 Oct;4(10):1677-86
references.pmid: 16215175
references.citation: Microbiology (Reading). 1997 Feb;143 ( Pt 2):331-340
references.pmid: 9043110
references.citation: Biochem J. 1990 Dec 1;272(2):407-14
references.pmid: 2268269
references.citation: Infect Immun. 2000 Dec;68(12):6712-9
references.pmid: 11083786
references.citation: Trends Cell Biol. 2010 Oct;20(10):580-5
references.pmid: 20685121
references.citation: Proc Natl Acad Sci U S A. 2009 Jan 13;106(2):599-604
references.pmid: 19116272
references.citation: Eukaryot Cell. 2005 Apr;4(4):633-8
references.pmid: 15821123
references.citation: Infect Control Hosp Epidemiol. 2005 Jun;26(6):540-7
references.pmid: 16018429
references.citation: J Biol Chem. 2002 Feb 8;277(6):3965-72
references.pmid: 11724795
references.citation: Microbiology (Reading). 2005 May;151(Pt 5):1645-1655
references.pmid: 15870472
references.citation: Cell. 1997 Sep 5;90(5):939-49
references.pmid: 9298905
references.citation: Physiol Rev. 1989 Jul;69(3):765-96
references.pmid: 2526342
references.citation: J Biol Chem. 2004 Aug 6;279(32):33131-8
references.pmid: 15180988
references.citation: Science. 1994 Dec 9;266(5191):1723-6
references.pmid: 7992058
references.citation: Genetics. 2002 Dec;162(4):1573-81
references.pmid: 12524333
references.citation: BMC Microbiol. 2010 Apr 30;10:133
references.pmid: 20433738
references.citation: Eukaryot Cell. 2010 Aug;9(8):1203-15
references.pmid: 20581294
references.citation: Microbiol Mol Biol Rev. 2006 Mar;70(1):177-91
references.pmid: 16524922
references.citation: J Biol Chem. 2008 Jul 18;283(29):20309-19
references.pmid: 18502746
references.citation: Proc Natl Acad Sci U S A. 2012 Feb 14;109(7):2234-9
references.pmid: 22308411
references.citation: Med Microbiol Immunol. 2013 Jun;202(3):183-95
references.pmid: 23354731
references.citation: Eukaryot Cell. 2010 Sep;9(9):1320-8
references.pmid: 20656912
references.citation: Fungal Genet Biol. 2008 Jun;45(6):861-77
references.pmid: 18296085
references.citation: Curr Biol. 1997 Nov 1;7(11):R691-4
references.pmid: 9382797
references.citation: Proc Natl Acad Sci U S A. 1988 Nov;85(21):7972-6
references.pmid: 2973058
references.citation: J Bioenerg Biomembr. 2005 Dec;37(6):431-5
references.pmid: 16691478
references.citation: Nature. 2012 Jan 09;481(7381):360-4
references.pmid: 22230956
references.citation: J Exp Biol. 2000 Jan;203(Pt 1):89-95
references.pmid: 10600677
references.citation: Proc Natl Acad Sci U S A. 1983 Mar;80(5):1300-3
references.pmid: 6131417
references.citation: J Biol Chem. 1990 Oct 25;265(30):18447-53
references.pmid: 2145283
references.citation: Mol Gen Genet. 1993 Nov;241(3-4):447-56
references.pmid: 8246899
references.citation: Nature. 2001 Feb 1;409(6820):581-8
references.pmid: 11214310
references.citation: J Bacteriol. 1993 Sep;175(17):5566-74
references.pmid: 8366041
references.citation: Methods Mol Med. 2005;118:71-9
references.pmid: 15888936
references.citation: Infect Control Hosp Epidemiol. 2008 Nov;29(11):996-1011
references.pmid: 18947320
references.citation: J Biol Chem. 2000 Dec 8;275(49):38245-53
references.pmid: 10991947
references.citation: J Gen Microbiol. 1988 May;134(5):1079-87
references.pmid: 3058860
references.citation: J Biol Chem. 1991 Jul 25;266(21):13971-7
references.pmid: 1830311
references.citation: Mol Cell Biol. 1998 Mar;18(3):1534-43
references.pmid: 9488470
references.citation: Nat Rev Microbiol. 2011 Aug 16;9(10):737-48
references.pmid: 21844880
references.citation: J Bacteriol. 1999 Mar;181(6):1868-74
references.pmid: 10074081
references.citation: Eukaryot Cell. 2012 May;11(5):683-93
references.pmid: 22427429
references.citation: Eukaryot Cell. 2006 Feb;5(2):411-21
references.pmid: 16467481
references.citation: Int J Cancer. 2013 Jan 15;132(2):E1-10
references.pmid: 22945374
references.citation: Yeast. 2000 Jan 15;16(1):65-70
references.pmid: 10620776
references.citation: Brief Bioinform. 2008 Jul;9(4):286-98
references.pmid: 18372315
references.citation: J Biol Chem. 2006 Oct 6;281(40):30001-14
references.pmid: 16891312
references.citation: Trends Microbiol. 2003 Jan;11(1):30-6
references.pmid: 12526852
references.citation: Mol Biol Cell. 2007 Oct;18(10):3873-82
references.pmid: 17652457
references.citation: J Exp Biol. 2009 Feb;212(Pt 3):341-6
references.pmid: 19151208
references.citation: PLoS Pathog. 2010 Jun 03;6(6):e1000939
references.pmid: 20532216