STUDY OF PHOSPHORYLATED RESIDUES OF LEPRK2 IN TOMATO AND THEIR FUNCTION IN POLLEN-PISTIL INTERACTION

SALEMTM; WENGIER D; MAZZELLA A; MUSCHIETTI J

Cordoba, Argentina

Congreso; SAIB; 2008

LePRK1 and LePRK2 are pollen-specific receptor kinases from Solanum lycopersicum, expressed in pollen tubes and potentially involved in pollen-pistil interactions. Transduction by their kinase activity of a pistil signal might initiate a signaling cascade in the tube that regulates its growth. LePRK2 is phosphorylated in vitro in pollen membranes. To know which LePRK2 residues are phosphorylated in vivo, we separated proteins from mature pollen by 2D gels. Potential phosphorylated isoforms were visualized with western blots against LePRK2 and isolated in silver stained 2D gels. Using a computational approach that predicts phosphorylation sites in LePRK2, we selected 8 serines or threonines of LePRK2: S277, S279, S282, S304, S307, T308, T358 and S396. To study their functional relevance in pollen tube growth, we substituted them by Alanine and Aspartic acid using for mutagenesis the pLAT52-LePRK2-eGFP construct, followed by transient expression in tobacco pollen. We previously showed pollen receiving LePRK2-eGFP displayed aberrant ballon tip morphology at germination. Pollen transfected with LePRK2(A277/A279/A282)-eGFP recovered the wt phenotype and are longer; neither of the constructs with isolated alanine residues showed this reversion. Contrary, pollen tubes receiving LePRK2(D277/D279/D282)-eGFP showed a wt phenotype. These results will be complemented by the proteomic approach.