Imaging the assembly, structure, and function of the nuclear pore inside cells.

The nuclear pore complex (NPC) mediates selective transport across the nuclear envelope (NE) and plays crucial roles in several additional cellular functions. In higher eukaryotes, the NPC and the NE disassemble and reassemble during cell division and live-cell imaging has been a powerful tool to analyze these dynamic processes. Here, we present a method for the kinetic analysis of postmitotic NPC assembly and reestablishment of transport competence in intact cells by multicolor 4D imaging and photoswitching. By applying the methods we have established previously using normal rat kidney to HeLa cells, we demonstrate the conservation of NPC assembly in different mammalian cells. We recently showed that the molecular organization of the NPC can be studied by combining stochastic super-resolution microscopy with single-particle averaging and present this method here in detail.