A STUDY OF ORM EXPRESSION CHANGES FUMONISIN B1-INDUCED PERTURBATIONS OF SPHINGOLIPID HOMEOSTASIS AND DIFFERENTIALLY TOUCHES CERAMIDE SYNTHASE ACTIVITIES

Abstract

Sphingolipid synthesis is tightly regulated in eukaryotes. This regulation in plants ensures sufcient sphingolipids to support growth, while limiting accumulation of sphingolipid metabolites that induce programmed cell death (PCD). Serine palmitoyltransfersase (SPT) catalyzes the rst step in sphingolipid biosynthesis and is considered the primary sphingolipid homeostatic regulatory point. In this report, Arabidopsis putative SPT regulatory proteins, orosomucoid-like proteins AtORM1 and AtORM2 were found to physically interact with the Arabidopsis SPT and to suppress SPT activity when co-expressed with Arabidopsis SPT subunits LCB1 and LCB2 and the small subunit of SPT in a yeast SPT-decient mutant. Consistent with a role in SPT suppression, AtORM1 and AtORM2 overexpression lines displayed increased resistance to the PCD-inducing mycotoxin fumonisin B1 (FB1), with an accompanying reduced accumulation of longchain bases (LCBs) and C16-fatty acid-containing ceramide accumulation relative to wild type plants. Conversely, RNAi suppression lines of AtORM1 and AtORM2 displayed increased sensitivity to FB1 and an accompanying strong increase in LCBs and C16 fatty acid-containing ceramides relative to wild-type plants. Overexpression lines were also found to have reduced activity of the Class I ceramide synthase that uses C16-fatty acid acyl-CoA and dihydroxy LCB substrates, but increased activity of Class II ceramide synthases that use very long-chain fatty acylCoA and trihydroxy LCB substrates. RNAi suppression lines, in contrast, displayed increased Class I ceramide synthase activity, but reduced Class II ceramide synthase activity. These ndings indicate that ORM-mediation of SPT activity differentially regulates functionally distinct ceramide synthase activities as part of a broader sphingolipid homeostatic regulatory network.