Sphingolipid remodeling in the plasma membrane is essential for osmotic stress tolerance in Arabidopsis

Osmotic stress caused by drought, salinity, or cold conditions is an important abiotic factor that decreases membrane integrity and causes cell death, thus decreasing plant growth and productivity. Remodeling cell membrane composition via lipid turnover can counter the loss of membrane integrity and cell death caused by osmotic stress. Sphingolipids are important components of eukaryotic membrane systems; however, how sphingolipids participate in plant responses to osmotic stress remains unclear. Here, we characterized the role of the glucosylceramidase (GCD) AtGCD1 (encoded by At1g33700) in sphingolipid remodeling and acclimation to osmotic stress in Arabidopsis (Arabidopsis thaliana). AtGCD1-AtGCD4 are Arabidopsis homologs of human nonlysosomal glucosylceramidase. We determined that AtGCD1 functions as a glucosylceramidase and localizes to the plasma membrane and that recombinant AtGCD1 has no substrate preference for acyl chain length. Moreover, AtGCD1 and AtGCD3 (At4g10060) are essential for osmotic stress tolerance in Arabidopsis. In cells treated with mannitol, AtGCD1 and AtGCD3 hydrolyzed glucosylceramides to ceramides, leading to decreased glucosylceramide contents and increased glycosyl inositol phosphoceramide contents. We observed a substantial change in the molecular order of lipids and membrane tension at the plasma membrane of the Arabidopsis gcd1 gcd3 double mutant, indicating that glucosylceramidases compensate for changes in membrane properties to stabilize the membrane during osmotic stress. Notably, we found that loss of GCD1 and GCD3 enhanced plant resistance to beet armyworm (Spodoptera exigua). Our results suggest that sphingolipid remodeling regulates the physicochemical properties of cellular membranes during plant stress responses.