Analysis of exocyst subunit EXO70 family reveals distinct membrane domains in tobacco pollen tubes

Sekereš J., Pejchar P., Šantrůček J., Vukasinović N., Žárský V., Potocký M.
PLANT PHYSIOLOGY 173(3): 1659-1675, 2017

Abstract: The vesicle-tethering complex exocyst is one of the crucial cell polarity regulators. EXO70 subunit is required for the targeting of the complex and is represented by many isoforms in angiosperm plant cells. This diversity could be partly responsible for establishment and maintenance of membrane domains with different composition. To address this hypothesis, we employed growing pollen tube, a well-established cell polarity model system, and performed large-scale expression, localization and functional analysis of tobacco EXO70 isoforms. Various isoforms localized to different regions of pollen tube plasma membrane, apical vesicle-rich inverted cone region, nucleus and cytoplasm. Overexpression of major pollen-expressed EXO70 isoforms resulted in growth arrest and characteristic phenotypic deviations of tip swelling and apical invaginations. NtEXO70A1a and NtEXO70B1 occupied two distinct and mutually exclusive plasma membrane domains. Both isoforms partly colocalized with exocyst subunit NtSEC3a at the plasma membrane, possibly forming different exocyst complex subpopulations. NtEXO70A1a localized to the small area previously characterized as the site of exocytosis in tobacco pollen tube, while NtEXO70B1 surprisingly colocalized with zone of clathrin-mediated endocytosis. Both NtEXO70A1a and NtEXO70B1 colocalized to a different degree with markers for anionic signaling phospholipids phosphatidylinositol 4,5-bisphosphate and phosphatidic acid. In contrast, members of EXO70 C-class, which are specifically expressed in tip growing cells, exhibited exocytosis-related functional effect in pollen tubes despite no apparent plasma membrane localization. Taken together, our data support the existence of multiple membrane-trafficking domains regulated by different EXO70-containing exocyst complexes within the single cell.
DOI: 10.​1104/​pp.​16.​01709 IEB authors: Přemysl Pejchar, Martin Potocký, Juraj Sekereš, Nemanja Vukasinovic, Viktor Žárský