Mutations in the Arabidopsis Phosphoinositide Phosphatase Gene SAC9 Lead to Overaccumulation of PtdIns(4,5)P2 and Constitutive Expression of the Stress-Response Pathway

Abstract

Phosphoinositides (PIs) are signaling molecules that regulate cellular events including vesicle targeting and interactions between membrane and cytoskeleton. Phosphatidylinositol (PtdIns)(4,5)P₂ is one of the best characterized PIs; studies in which PtdIns(4,5)P₂ localization or concentration is altered lead to defects in the actin cytoskeleton and exocytosis. PtdIns(4,5)P₂ and its derivative Ins(1,4,5)P₃ accumulate in salt, cold, and osmotically stressed plants. PtdIns(4,5)P₂ signaling is terminated through the action of inositol polyphosphate phosphatases and PI phosphatases including supressor of actin mutation (SAC) domain phosphatases. In some cases, these phosphatases also act on Ins(1,4,5)P₃. We have characterized the Arabidopsis (Arabidopsis thaliana) sac9 mutants. The SAC9 protein is different from other SAC domain proteins in several ways including the presence of a WW protein interaction domain within the SAC domain. The rice (Oryza sativa) and Arabidopsis SAC9 protein sequences are similar, but no apparent homologs are found in nonplant genomes. High-performance liquid chromatography studies show that unstressed sac9 mutants accumulate elevated levels of PtdIns(4,5)P₂ and Ins(1,4,5)P₃ as compared to wild-type plants. The sac9 mutants have characteristics of a constitutive stress response, including dwarfism, closed stomata, and anthocyanin accumulation, and they overexpress stress-induced genes and overaccumulate reactive-oxygen species. These results suggest that the SAC9 phosphatase is involved in modulating phosphoinsitide signals during the stress response.