Dataset Information

A mitogen-activated protein kinase limits the apical formation of phosphatidylinositol 4,5-bisphosphate during polarized cell expansion of pollen tubes

ABSTRACT: Mitogen activated protein kinases (MAPKs) and phosphoinositides contribute to establishment and maintenance of polarity in eukaryotic cells. However, interactions between these important signaling pathways are largely unclear. Pollen tubes are an important model for the study of polarized cell expansion, in which PI4P 5-kinases control apical secretion and MAPKs contribute to pollen tube guidance. Here we report on the regulation of the Arabidopsis PI4P 5-kinase, AtPIP5K6, by the MAPK, MPK6. Purified recombinant AtPIP5K6 was phosphorylated in vitro upon incubation with pollen tube extracts. Non-targeted in-gel-kinase assays of pollen tube extracts followed by mass-spectrometry identified MPK6 as a candidate protein kinase upstream of AtPIP5K6. MPK6 interacted with AtPIP5K6 in yeast-two-hybrid tests, and bimolecular fluorescence complementation verified the interaction at the apical plasma membrane of pollen tubes. Recombinant AtPIP5K6 was phosphorylated in vitro by recombinant MPK6, resulting in reduced catalytic activity of AtPIP5K6. Alanine-substitution of phosphorylation sites, T590A and T597A, abrogated phosphorylation of AtPIP5K6 by MPK6 and prevented the in vitro inhibition. A T597D substitution resulted in reduced catalytic activity of the modified AtPIP5K6 protein in vitro. Coexpression of MPK6 with AtPIP5K6 in pollen tubes resulted in attenuation of PI4P 5-kinase-mediated polarity defects in vivo. Reciprocally, AtPIP5K6 effects were enhanced upon concomitant RNAi-inhibition of endogenous tobacco MAPKs. Both effects were absent when a AtPIP5K6 T590A T597A-variant was expressed, which could no longer be phosphorylated. We conclude that MPK6 limits PI4P 5-kinase-dependent polarized secretion to effect pollen tube guidance, revealing a new link between conserved signaling pathways with potential ramifications across eukaryotic kingdoms.

INSTRUMENT(S): LTQ Orbitrap Velos, Synapt MS

ORGANISM(S): Nicotiana Tabacum (common Tobacco) Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Plant Cell, Pollen

SUBMITTER: Sacha Baginsky

LAB HEAD: Ingo Heilmann

PROVIDER: PXD006067 | Pride | 2022-02-22

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	FH010713_2_2_MPK6_target.msf.zip	Msf
	FH010713_2_2_MPK6_target.raw.zip	Raw
	FH260813_PIPK6_T590A_MPK6_DDA.msf.zip	Msf
	FH260813_PIPK6_T590A_MPK6_DDA.raw.zip	Raw
	FH260813_PIPK6_T590A_MPK6_DIA.msf.zip	Msf

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Publications

MAPKs Influence Pollen Tube Growth by Controlling the Formation of Phosphatidylinositol 4,5-Bisphosphate in an Apical Plasma Membrane Domain.

Hempel Franziska F Stenzel Irene I Heilmann Mareike M Krishnamoorthy Praveen P Menzel Wilhelm W Golbik Ralph R Helm Stefan S Dobritzsch Dirk D Baginsky Sacha S Lee Justin J Hoehenwarter Wolfgang W Heilmann Ingo I

The Plant cell 20171122 12

An apical plasma membrane domain enriched in the regulatory phospholipid phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P<sub>2</sub>] is critical for polar tip growth of pollen tubes. How the biosynthesis of PtdIns(4,5)P<sub>2</sub> by phosphatidylinositol 4-phosphate 5-kinases (PI4P 5-kinases) is controlled by upstream signaling is currently unknown. The pollen-expressed PI4P 5-kinase PIP5K6 is required for clathrin-mediated endocytosis and polar tip growth in pollen tubes. Here, we identif ...[more]

PMID: 29167320

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