Permeability of Single Nuclear Pores

Abstract

In this first application of optical single transporter recording (OSTR), a recently established technique for optically monitoring the activity of single transporters in membrane patches (Tschödrich-Rotter and Peters. 1998. J. Microsc. 192:114–125), the passive permeability of the nuclear pore complex (NPC) was measured for a homologous series of hydrophilic probe molecules. Nuclei were isolated from Xenopus oocytes and firmly attached to filters containing small cylindrical pores. Transport through membrane patches spanning filter pores was measured by scanning microphotolysis. Thus the permeability coefficients of single NPCs were determined for fluorescently labeled dextrans of ∼4, 10, and 20kDa. Dextrans of ≥40kDa could not permeate the NPC. The data were consistent with a model in which the NPC contains a single diffusion channel. By application of established theories for the restricted diffusion through small pores, the diffusion channel was approximated as a cylinder with a radius of 4.4–6.1nm (mean 5.35nm). Because the transport rate constant of the single NPC was known, the equivalent length of the channel could be also determined and was found to be 40–50nm (mean 44.5nm). The symmetry of the NPC implies that a singular component such as the diffusion channel is located at the center of the NPC. Therefore a common transport pathway apparently mediates both passive and signal-dependent transport. To test this hypothesis, measurements of signal-dependent transport and of the mutual effects signal-dependent and passive transport may exert on each other are in progress.