Cortical actin contributes to spatial organization of ER–PM junctions

Endoplasmic reticulum-plasma membrane (ER-PM) junctions mediate crucial activities ranging from Ca2+ signaling to lipid metabolism. Spatial organization of ER-PM junctions may modulate the extent and location of these cellular activities. However, the morphology and distribution of ER-PM junctions are not well characterized. Using photoactivated localization microscopy, we reveal that the contact area of single ER-PM junctions is mainly oblong with the dimensions of ∼120 nm × ∼80 nm in HeLa cells. Using total internal reflection fluorescence microscopy and structure illumination microscopy, we show that cortical actin contributes to spatial distribution and stability of ER-PM junctions. Further functional assays suggest that intact F-actin architecture is required for phosphatidylinositol 4,5-bisphosphate homeostasis mediated by Nir2 at ER-PM junctions. Together, our study provides quantitative information on spatial organization of ER-PM junctions that is in part regulated by F-actin. We envision that functions of ER-PM junctions can be differentially regulated through dynamic actin remodeling during cellular processes.