Mutations in Two Putative Phosphorylation Motifs in the Tomato Pollen Receptor Kinase LePRK2 Show Antagonistic Effects on Pollen Tube Length

SALEM, TAMARA; MAZZELLA, AGUSTINA; BARBERINI, MARIA LAURA; WENGIER, DIEGO; MOTILLO, VIVIANA; PARISI, GUSTAVO; MUSCHIETTI, JORGE

JOURNAL OF BIOLOGICAL CHEMISTRY (ONLINE)

ASBMB

Año: 2011 vol. 286 p. 4882 - 4891

1083-351X

The tip-growing pollen tube is a useful model for studyingpolarized cell growth in plants. We previously characterizedLePRK2, a pollen-specific receptor-like kinase from tomato(1). Here, we showed that LePRK2 is present as multiplephosphorylated isoforms in mature pollen membranes. Usingcomparative sequence analysis and phosphorylation siteprediction programs, we identified two putative phosphorylationmotifs in the cytoplasmic juxtamembrane (JM) domain.Site-directed mutagenesis in these motifs, followed bytransient overexpression in tobacco pollen, showed thatboth motifs have opposite effects in regulating pollen tubelength. Relative to LePRK2-eGFP pollen tubes, alanine substitutionsin residues of motif I, Ser277/Ser279/Ser282, resultedin longer pollen tubes, but alanine substitutions inmotif II, Ser304/Ser307/Thr308, resulted in shorter tubes. Incontrast, phosphomimicking aspartic substitutions at theseresidues gave reciprocal results, that is, shorter tubes withmutations in motif I and longer tubes with mutations inmotif II. We conclude that the length of pollen tubes can benegatively and positively regulated by phosphorylation ofresidues in motif I and II respectively. We also showed thatLePRK2-eGFP significantly decreased pollen tube lengthand increased pollen tube tip width, relative to eGFP tubes.The kinase activity of LePRK2 was relevant for this phenotypebecause tubes that expressed a mutation in a lysine essentialfor kinase activity showed the same length and widthas the eGFP control. Taken together, these results suggestthat LePRK2 may have a central role in pollen tube growththrough regulation of its own phosphorylation status.