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  • Unregulated Sphingolipid Biosynthesis in Gene-Edited Arabidopsis ORM Mutants Results in Nonviable Seeds with Strongly Reduced Oil Content
Title: Unregulated Sphingolipid Biosynthesis in Gene-Edited Arabidopsis ORM Mutants Results in Nonviable Seeds with Strongly Reduced Oil Content
Orosomucoid-like proteins (ORMs) interact with serine palmitoyltransferase (SPT) to negatively regulate sphingolipid biosynthesis, a reversible process critical for balancing the intracellular sphingolipid levels needed for growth and programmed cell death. Here we show that ORM1 and ORM2 are essential for lifecycle completion in Arabidopsis thaliana. Seeds from orm1−/− orm2−/− mutants (generated by crossing CRISPR/Cas9 knockout mutants for each gene) accumulated high levels of ceramide, pointing to unregulated sphingolipid biosynthesis. orm1−/− orm2−/− seeds were nonviable, displayed aberrant embryo development, and had >80% reduced oil content vs. wild-type seeds. This phenotype was mimicked in Arabidopsis seeds expressing the SPT subunit LCB1 lacking its first transmembrane domain, which is critical for ORM-mediated regulation of SPT. We identified a mutant for ORM1 lacking one amino acid (Met51) near its second transmembrane domain that retained its membrane topology. Expressing this allele in the orm2 background yielded plants that did not advance beyond the seedling stage, hyperaccumulated ceramides, and showed altered organellar structures and increased senescence and pathogenesis-related gene expression. These seedlings also showed upregulated expression of genes for sphingolipid catabolic enzymes, pointing to additional mechanisms for maintaining sphingolipid homeostasis. ORM1 lacking Met51 had strongly impaired interactions with LCB1 in yeast (Saccharomyces cerevisiae) model, providing structural clues about regulatory interactions between ORM and SPT.  more » « less
Award ID(s):
1818297
PAR ID:
10173426
Author(s) / Creator(s):
; ; ; ; ; ; ;
Date Published:
Journal Name:
The Plant Cell
ISSN:
1040-4651
Page Range / eLocation ID:
tpc.00015.2020
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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