Poster: Intracellular Signaling

Abs # 839: Characterization of Arabidopsis thaliana inositol 5-phosphatases: phosphoinositide signal transduction terminators

Presenter:	Burnette, Ryan N, rburnett@vt.edu
Authors	Burnette, Ryan N (A)   Ercetin, Mustafa  (A)   Gillaspy, Glenda  (A)
Affiliations:	(A): Virginia Tech

Plants require diverse signaling pathways to mediate environmental changes. There is growing evidence that plants use phosphoinositides (PI)s as components of certain signaling pathways. Inositol 1,4,5-trisphosphate has been implicated as a second messenger in plants, notably in abscisic acid (ABA) signal transduction. The Arabidopsis genome encodes 15 inositol 5-phosphatases (At5PTases) capable of removing a 5-phosphate from a variety of substrates: I(1,4,5)P₃, I(1,3,4,5)P₄, PI(4,5)P₂ and PI(3,4,5)P₃. We have previously shown that At5PTase1 has the ability to hydrolyze a 5-phosphate from I(1,4,5)P₃ and I(1,3,4,5)P₄. Ectopic expression of At5PTase1 interferes with ABA signaling by reducing I(1,4,5)P₃ levels, resulting in ABA-insensitive stomata. To examine the substrate selectivity of the At5PTases, recombinant At5PTase proteins were expressed in Drosophila S2 cells. Immunoprecipitated recombinant At5PTases were analyzed in activity assays. We have investigated the catalytic activity of 3 different At5PTases which contain diverse sequences outside of the catalytic domain. In addition to hydrolyzing a 5-phosphate from I(1,4,5)P₃ and I(1,3,4,5)P₄, recombinant At5PTase1 also catalyzes the removal of a phosphate from the lipid substrates PI(4,5)P₂ and PI(3,4,5)P₃ in vitro. In contrast, At5PTase11 only hydrolyzes the lipid substrates PI(4,5)P₂ and PI(3,4,5)P₃ in vitro. These findings indicate that the At5PTases discriminate between different IP and PIP substrates, and may function in different signaling pathways. Another member of the At5PTases, At5PTase12, has several WD domain repeats in the amino-terminus. These domains have been implicated in protein-protein interactions. Recombinant At5PTase12 is currently being investigated to determine its substrate specificity.