Cortical actin contributes to spatial organization of ER–PM junctions

Endoplasmic reticulum–plasma membrane (ER–PM) junctions mediate crucial activities ranging from Ca(2+) 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.