The class VIII myosin ATM2 stabilizes actin-plasma membrane contacts by locally promoting PtdIns(4,5)P2 nanodomain formation in tobacco pollen tubes

Abstract The intersection of actin-filaments with the plasma membrane in plant cells involves lipid-protein interactions by the Arabidopsis class VIII-myosin, ATM2 that are currently not understood. Using pollen tube cells as a model, we describe how ATM2 enables actin-plasma membrane contacts by modulating membrane lipid nano-organization. ATM2 binds both actin and phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2), a lipid required for ATM2-plasma membrane-attachment. Overexpression of ATM2 unexpectedly stabilized actin-dynamics and altered pollen tube-morphology. Actin-dynamics remained unaltered upon overexpression of a lipid binding-defective ATM2-variant. ATM2-effects on actin-dynamics required PtdIns(4,5)P2, as artificial PtdIns(4,5)P2-depletion in pollen tubes abolished effects of full-length ATM2 on actin dynamics despite its retained plasma membrane-association. ATM2 colocalized and interacted with the PI4P 5-kinase, PIP5K2, which resides at actin-plasma membrane-contacts and forms PtdIns(4,5)P2-nanodomains facilitating ROP-dependent actin-stabilization. Upon ATM2-overexpression, a fluorescent PtdIns(4,5)P2-biosensor decorated an expanded plasma membrane-region in vivo, indicating a promoting effect of ATM2 on PtdIns(4,5)P2-abundance. Moreover, catalytic activity of purified recombinant PIP5K2 protein was enhanced upon coincubation with a purified C-terminal ATM2-fragment in vitro, suggesting that ATM2 contributes to the intrinsic regulation of PtdIns(4,5)P2-formation. Actin-dynamics are, thus, stabilized at the actin-plasma membrane interface by ATM2 locally promoting the formation of PtdIns(4,5)P2-nanodomains, thereby self-reinforcing ATM2-recruitment while also facilitating ROP-activation to stabilize membrane-proximal actin-filaments.