Arabidopsis phosphatidylinositol-4-monophosphate 5-kinase 4 regulates pollen tube growth and polarity by modulating membrane recycling.

Abstract

Phosphatidylinositol-4-monophosphate 5-kinases produce phosphatidylinositol (4,5)-bisphosphate [PtdIns(4,5)P(2)] and have been implicated in vesicle trafficking and cytoskeletal rearrangements. Here, we adopted a reverse genetics approach to investigate the function of the Arabidopsis thaliana pollen-expressed gene encoding phosphatidylinositol-4-monophosphate 5-kinase 4 (PIP5K4). Pollen germination, tube growth, and polarity were significantly impaired in homozygous mutant plants lacking PIP5K4 transcript. In vitro, supplementation with PtdIns(4,5)P(2) rescued these phenotypes. In vivo, mutant pollen fertilized ovules, leading to normal seed set and silique length. However, fertilization took longer than in wild-type plants, and the pip5k4 null mutant allele was transmitted through the pollen at a reduced frequency. Analysis of endocytic events using FM1-43 (or FM4-64) suggested a reduction in endocytosis and membrane recycling in pip5k4 null mutant pollen tubes. Imaging of elongating tobacco (Nicotiana tabacum) pollen tubes transiently transformed with a PIP5K4-green fluorescent protein fusion construct revealed that the protein localized to the plasma membrane, particularly in the subapical region. Overexpression of PIP5K4-GFP delocalized the protein to the apical region of the plasma membrane, perturbed pollen tube growth, and caused apical cell wall thickening. Thus, PIP5K4 plays a crucial role in regulating the polarity of pollen tubes. This study supports a model for membrane secretion and recycling where the apical and subapical regions appear to contain the components required to promote and sustain growth.