Phosphoinositides Regulate Clathrin-Dependent Endocytosis at the Tip of Pollen Tubes inArabidopsisand Tobacco

Abstract

Using the tip-growing pollen tube of Arabidopsis thaliana and Nicotiana tabacum as a model to investigate endocytosis mechanisms, we show that phosphatidylinositol-4-phosphate 5-kinase 6 (PIP5K6) regulates clathrin-dependent endocytosis in pollen tubes. Green fluorescent protein-tagged PIP5K6 was preferentially localized to the subapical plasma membrane (PM) in pollen tubes where it apparently converts phosphatidylinositol 4-phosphate (PI4P) to phosphatidylinositol 4,5-bisphosphate [PI(4,5)P 2 ]. RNA interference-induced suppression of PIP5K6 expression impaired tip growth and inhibited clathrin-dependent endocytosis in pollen tubes. By contrast, PIP5K6 overexpression induced massive aggregation of the PM in pollen tube tips. This PM abnormality was apparently due to excessive clathrin-dependent membrane invagination because this defect was suppressed by the expression of a dominant-negative mutant of clathrin heavy chain. These results support a role for PI(4,5)P 2 in promoting early stages of clathrin-dependent endocytosis (i.e., membrane invagination). Interestingly, the PIP5K6 overexpression-induced PM abnormality was partially suppressed not only by the overexpression of PLC2, which breaks down PI(4,5)P 2 , but also by that of PI4Kb1, which increases the pool of PI4P. Based on these observations, we propose that a proper balance between PI4P and PI(4,5)P 2 is required for clathrin-dependent endocytosis in the tip of pollen tubes.

Arabidopsis phosphatidylinositol monophosphate 5-kinase 2 is involved in root gravitropism through regulation of polar auxin transport by affecting the cycling of PIN proteins

“…In addition, exogenous PtdIns(4,5)P 2 stimulates the internalization process in WT cells ( Figure 5B), which strongly suggests that vesicle trafficking, especially endocytosis, is affected by PIP5K2 through its product PtdIns(4,5)P 2 . These results are similar to previous findings that disruption of PIP5K4 inhibits proper endocytosis and membrane recycling in pollen tubes [21] and consistent with the report that PIP5K6 and PtdIns(4,5)P 2 positively regulate clathrin-dependent endocytosis [18].…”

“…Besides 4 homologs of yeast PIP5K, 11 PIPK isoforms can be divided into two subfamilies (A and B) according to their domain organization [14]. The catalytic activities of using PtdIns4P as the preferred substrate to generate PtdIns(4,5)P 2 have all been characterized [15][16][17][18]. Studies have revealed the physiological functions of some PIP5Ks, including PIP5K1 in abscisic acid signaling [19]; PIP5K3 in root hair formation and growth [16,20]; PIP5K4 in stomata opening [21]; and PIP5K4-6 in pollen tube growth [17,22].…”

Tissue-specific expression, developmentally and spatially regulated alternative splicing, and protein subcellular localization of OsLpa1 in rice

“…2g), suggesting that it may play an essential role, possibly in filament elongation at anthesis Zhao et al, 2010). This would not be surprising given that two phosphatidylinositol compounds in the lipiddependent PA synthesis pathway, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P 2 ) and phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P 3 ), participate in the recruitment and activation of a wide variety of actin regulatory proteins at the plasma membrane, controlling cell shape, motility, and cytokinesis (di Paolo and de Camilli, 2006).…”

CASEIN KINASE1-LIKE PROTEIN2 Regulates Actin Filament Stability and Stomatal Closure via Phosphorylation of Actin Depolymerizing Factor

“…For instance, actin dynamics have been implicated in mediating Ca 2+ concentration changes in the cytosol, K + channel activity, and vacuole shape determination (Hwang et al, 1997;Gao et al, 2009;Zhao et al, 2013), but the related molecular mechanisms remain to be established. One possibility is regulation of membrane recycling by actin reorganization Lee et al, 2008;Zhao et al, 2010;Bou Daher and Geitmann, 2011;Zhu et al, 2013), which would change the localization of ion channels/transporters and the area of vacuolar membrane. Actin dynamics may also act as an important component in signal transduction to mediate such changes (Gao et al, 2008;Zhao et al, 2011Zhao et al, , 2013Jiang et al, 2012).…”