A gene encoding a sphingolipid biosynthesis enzyme determines the sensitivity of Saccharomyces cerevisiae to an antifungal plant defensin from dahlia (Dahlia merckii).
We have previously identified a Saccharomyces cerevisiae mutant that is markedly more resistant than wild-type to Dahlia merckii antimicrobial peptide 1 (DmAMP1), an antifungal plant defensin isolated from seeds of dahlia (Dahlia merckii). A complementation approach was followed that consisted of the introduction of a genomic library of DmAMP1-sensitive wild-type yeast into the DmAMP1-resistant yeast mutant and screening for restored sensitivity to DmAMP1. The gene determining sensitivity of S. cerevisiae to DmAMP1 was identified as IPT1, a gene encoding an enzyme involved in the last step of the synthesis of the sphingolipid mannose-(inositol-phosphate)(2)-ceramide. Strains with a nonfunctional IPT1 allele lacked mannose-(inositol-phosphate)(2)-ceramide in their plasma membranes, bound significantly less DmAMP1 compared with wild-type strains, and were highly resistant to DmAMP1-mediated membrane permeabilization. All of these phenotypic deviations could be restored by reintroduction of a functional IPT1 gene. Our data support a model in which membrane patches containing sphingolipids act as binding sites for DmAMP1 or, alternatively, are required to anchor membrane or cell wall-associated proteins, which themselves interact with DmAMP1.
F. A. Janssens Laboratory of Genetics, Katholieke Universiteit Leuven, Belgium.
K
K
Thevissen
B P
BP
Cammue
K
K
Lemaire
J
J
Winderickx
R C
RC
Dickson
R L
RL
Lester
K K
KK
Ferket
F
F
Van Even
A H
AH
Parret
W F
WF
Broekaert
D000483:Alleles; D000935:Antifungal Agents / Q000378:metabolism / Q000494:pharmacology*; D019659:Asteraceae / Q000737:chemistry*; D001665:Binding Sites; D002455:Cell Division / Q000187:drug effects; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects; D003001:Cloning, Molecular; D023082:Defensins; D005800:Genes, Fungal / Q000235:genetics; D005816:Genetic Complementation Test; D008826:Microbial Sensitivity Tests; D009154:Mutation / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism / Q000494:pharmacology*; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D013107:Sphingolipids / Q000096:biosynthesis* / Q000378:metabolism
D000935:Antifungal Agents; D023082:Defensins; C099969:Dm-AMP1 protein, Dahlia merckii; D010940:Plant Proteins; D029701:Saccharomyces cerevisiae Proteins; D013107:Sphingolipids; C109311:IPT1 protein, S cerevisiae; D017853:Phosphotransferases (Alcohol Group Acceptor)