Candida albicans sphingolipid C9-methyltransferase is involved in hyphal elongation

C9-methylated glucosylceramide is a fungus-specific sphingolipid. This lipid is a major membrane component in the cell and is thought to play important roles in the growth and virulence of several fungal species. To investigate the importance of the methyl branch of the long-chain base in glucosylceramides in pathogenic fungi, we identified and characterized a sphingolipid C9-methyltransferase gene (MTS1, C9-MethylTransferase for Sphingolipid 1) in the pathogenic yeast Candida albicans. The mts1 disruptant lacked (E,E)-9-methylsphinga-4,8-dienine in its glucosylceramides and contained (E)-sphing-4-enine and (E,E)-sphinga-4,8-dienine. Reintroducing the MTS1 gene into the mts1 disruptant restored the synthesis of (E,E)-9-methylsphinga-4,8-dienine in the glucosylceramides. We also created a disruptant of the HSX11 gene, encoding glucosylceramide synthase, which catalyses the final step of glucosylceramide synthesis, in C. albicans and compared this mutant with the mts1 disruptant. The C. albicans mts1 and hsx11 disruptants both had a decreased hyphal growth rate compared to the wild-type strain. The hsx11 disruptant showed increased susceptibility to SDS and fluconazole, similar to a previously reported sld1 disruptant that contained only (E)-sphing-4-enine in its glucosylceramides, suggesting that these strains have defects in their cell membrane structures. In contrast, the mts1 disruptant grew similarly to wild-type in medium containing SDS or fluconazole. These results suggest that the C9-methyl group of a long-chain base in glucosylceramides plays an important role in the hyphal elongation of C. albicans independent of lipid membrane disruption.

Download Full-text

Sphingolipids Mediate Differential Echinocandin Susceptibility in Candida albicans and Aspergillus nidulans

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.04667-14 ◽

2015 ◽

Vol 59 (6) ◽

pp. 3377-3384 ◽

Cited By ~ 9

Author(s):

Kelley R. Healey ◽

Krishna K. Challa ◽

Thomas D. Edlind ◽

Santosh K. Katiyar

Keyword(s):

Candida Albicans ◽

Aspergillus Nidulans ◽

Laboratory Strain ◽

Wild Type ◽

Enoyl Reductase ◽

Content Type ◽

Long Chain Base ◽

Membrane Target ◽

Sphingolipid Biosynthesis ◽

Long Chain

ABSTRACTThe cell wall synthesis-inhibiting echinocandins, including caspofungin and micafungin, play important roles in the treatment of candidiasis and aspergillosis. Previous studies revealed that, in the haploid yeastCandida glabrata, sphingolipid biosynthesis pathway mutations confer caspofungin reduced susceptibility (CRS) but micafungin increased susceptibility (MIS). Here, we describe oneCandida albicansstrain (of 10 tested) that similarly yields CRS-MIS mutants at relatively high frequency. Mutants demonstrated increased levels of long-chain bases (sphingolipid pathway intermediates) and, unique to this strain, loss of His104/Pro104 heterozygosity in theTSC13-encoded enoyl reductase. CRS-MIS was similarly observed in aC. albicanshomozygousfen1Δfen12Δ laboratory strain and in diverse wild-type strains following exogenous long-chain-base treatment. Analogous to these results, CRS-MIS was demonstrated in anAspergillus nidulansbasAmutant encoding defective sphingolipid C4-hydroxylase and in its wild-type parent exposed to long-chain bases. Sphingolipids likely modulate echinocandin interaction with their Fks membrane target in all susceptible fungi, with potential implications for optimizing therapy with existing antifungals and the development of novel agents.

Download Full-text

Ceramide/Long-Chain Base Phosphate Rheostat in Saccharomyces cerevisiae: Regulation of Ceramide Synthesis by Elo3p and Cka2p

Eukaryotic Cell ◽

10.1128/ec.2.2.284-294.2003 ◽

2003 ◽

Vol 2 (2) ◽

pp. 284-294 ◽

Cited By ~ 51

Author(s):

Scott D. Kobayashi ◽

Marek M. Nagiec

Keyword(s):

Saccharomyces Cerevisiae ◽

Protein Kinase ◽

Wild Type ◽

Α Subunit ◽

Chain Base ◽

Long Chain Base ◽

Ceramide Synthesis ◽

Long Chain ◽

Ck2 Kinase

ABSTRACT Sphingolipid precursors, namely, ceramide and long-chain base phosphates (LCBPs), are important growth regulators with often opposite effects on mammalian cells. A set of enzymes that regulate the levels of these precursors, referred to as a ceramide/LCBP rheostat, is conserved in all eukaryotes. In order to gain further insight into the function of the rheostat in Saccharomyces cerevisiae, we searched for mutants that are synthetically lethal with a deletion of the LCB3 gene encoding LCBP phosphatase. In addition to acquiring expected mutants lacking the LCBP lyase, the screen revealed elo3 (sur4) mutants that were defective in fatty acid elongation and cka2 mutants lacking the α′ subunit of the protein kinase CK2 (casein kinase). Both mutations affected the in vivo activity of the acyl coenzyme A (acyl-CoA)-dependent and fumonisin B1-sensitive ceramide synthase (CS). The Elo3 protein is necessary for synthesis of C26-CoA, which in wild-type yeast is a source of C26 fatty acyls found in the ceramide moieties of all sphingolipids. In the in vitro assay, CS had a strong preference for acyl-CoAs containing longer acyl chains. This finding suggests that a block in the formation of C26-CoA in yeast may cause a reduction in the conversion of LCBs into ceramides and lead to an overaccumulation of LCBPs that is lethal in strains lacking the Lcb3 phosphatase. In fact, elo3 mutants were found to accumulate high levels of LCBs and LCBPs. The cka2 mutants, on the other hand, exhibited only 25 to 30% of the in vitro CS activity found in wild-type membranes, indicating that the α′ subunit of CK2 kinase is necessary for full activation of CS. The cka2 mutants also accumulated high levels of LCBs and had elevated levels of LCBPs. In addition, both the elo3 and cka2 mutants showed increased sensitivity to the CS inhibitors australifungin and fumonisin B1. Together, our data demonstrate that the levels of LCBPs in yeast are regulated by the rate of ceramide synthesis, which depends on CK2 kinase activity and is also strongly affected by the supply of C26-CoA. This is the first evidence indicating the involvement of protein kinase in the regulation of de novo sphingolipid synthesis in any organism.

Download Full-text

RTA2 is involved in calcineurin-mediated azole resistance and sphingoid long-chain base release in Candida albicans

Cellular and Molecular Life Sciences ◽

10.1007/s00018-008-8409-3 ◽

2008 ◽

Vol 66 (1) ◽

pp. 122-134 ◽

Cited By ~ 28

Author(s):

X. M. Jia ◽

Y. Wang ◽

Y. Jia ◽

P. H. Gao ◽

Y. G. Xu ◽

...

Keyword(s):

Candida Albicans ◽

Azole Resistance ◽

Chain Base ◽

Long Chain Base ◽

Long Chain

Download Full-text

Catalase gene disruptant of the human pathogenic yeast Candida albicans is defective in hyphal growth, and a catalase-specific inhibitor can suppress hyphal growth of wild-type cells

Microbiology and Immunology ◽

10.1111/j.1348-0421.2008.00006.x ◽

2008 ◽

Vol 52 (1) ◽

pp. 16-24 ◽

Cited By ~ 10

Author(s):

Yoshiyuki Nakagawa

Keyword(s):

Candida Albicans ◽

Specific Inhibitor ◽

Hyphal Growth ◽

Wild Type ◽

Pathogenic Yeast ◽

Catalase Gene

Download Full-text

Enzymology of long-chain base synthesis by aorta: induction of serine palmitoyltransferase activity in rabbit aorta during atherogenesis.

The Journal of Lipid Research ◽

10.1016/s0022-2275(20)38793-9 ◽

1988 ◽

Vol 27 (7) ◽

pp. 763-770

Author(s):

R D Williams ◽

D S Sgoutas ◽

G S Zaatari

Keyword(s):

Rabbit Aorta ◽

Serine Palmitoyltransferase ◽

Chain Base ◽

Long Chain Base ◽

Long Chain

Download Full-text

The DNA Binding Protein Rfg1 Is a Repressor of Filamentation inCandida albicans

Genetics ◽

10.1093/genetics/157.4.1503 ◽

2001 ◽

Vol 157 (4) ◽

pp. 1503-1512 ◽

Cited By ~ 1

Author(s):

Roy A Khalaf ◽

Richard S Zitomer

Keyword(s):

Dna Binding ◽

Binding Protein ◽

Hyphal Growth ◽

Dna Binding Protein ◽

Homozygous Deletion ◽

Wild Type ◽

Pathogenic Yeast ◽

Repression Complex ◽

Type Gene ◽

Repression Domain

AbstractWe have identified a repressor of hyphal growth in the pathogenic yeast Candida albicans. The gene was originally cloned in an attempt to characterize the homologue of the Saccharomyces cerevisiae Rox1, a repressor of hypoxic genes. Rox1 is an HMG-domain, DNA binding protein with a repression domain that recruits the Tup1/Ssn6 general repression complex to achieve repression. The C. albicans clone also encoded an HMG protein that was capable of repression of a hypoxic gene in a S. cerevisiae rox1 deletion strain. Gel retardation experiments using the purified HMG domain of this protein demonstrated that it was capable of binding specifically to a S. cerevisiae hypoxic operator DNA sequence. These data seemed to indicate that this gene encoded a hypoxic repressor. However, surprisingly, when a homozygous deletion was generated in C. albicans, the cells became constitutive for hyphal growth. This phenotype was rescued by the reintroduction of the wild-type gene on a plasmid, proving that the hyphal growth phenotype was due to the deletion and not a secondary mutation. Furthermore, oxygen repression of the hypoxic HEM13 gene was not affected by the deletion nor was this putative ROX1 gene regulated positively by oxygen as is the case for the S. cerevisiae gene. All these data indicate that this gene, now designated RFG1 for Repressor of Filamentous Growth, is a repressor of genes required for hyphal growth and not a hypoxic repressor.

Download Full-text