Visualizing Intracellular Organelle and Cytoskeletal Interactions at Nanoscale Resolution on Millisecond Timescales.

Summary In eukaryotic cells, organelles and the cytoskeleton undergo highly dynamic yet organized interactions capable of orchestrating complex cellular functions. Visualizing these interactions requires noninvasive, long-duration imaging of the intracellular environment at high spatiotemporal resolution and low background. To achieve these normally opposing goals, we developed grazing incidence structured illumination microscopy (GI-SIM) that is capable of imaging dynamic events near the basal cell cortex at 97-nm resolution and 266 frames/s over thousands of time points. We employed multi-color GI-SIM to characterize the fast dynamic interactions of diverse organelles and the cytoskeleton, shedding new light on the complex behaviors of these structures. Precise measurements of microtubule growth or shrinkage events helped distinguish among models of microtubule dynamic instability. Analysis of endoplasmic reticulum (ER) interactions with other organelles or microtubules uncovered new ER remodeling mechanisms, such as hitchhiking of the ER on motile organelles. Finally, ER-mitochondria contact sites were found to promote both mitochondrial fission and fusion. Graphical Abstract Highlights • Super-resolution live-cell imaging up to 266 fps at 97-nm resolution • Hitchhiking interactions among organelles remodel ER and mitochondrial networks • ER-mitochondrion contacts promote coalescence of mitochondrial membranes • Collision of late endosomes or lysosomes carried along microtubules split ER tubules A new approach for visualizing dynamic processes within cells offers insight into membrane-membrane contact interactions and microtubule function. [ABSTRACT FROM AUTHOR]