ATP-INDUCED SHAPE CHANGE OF NUCLEAR-PORES VISUALIZED WITH THE ATOMIC-FORCE MICROSCOPE

Bidirectional transport of molecules between nucleus and cytoplasm thr ough the nuclear pore complexes (NPCs) spanning the nuclear envelope p lays a fundamental role in cell function and metabolism. Nuclear impor t of macromolecules is a two-step process involving initial recognitio n of targeting signals, docking to the pore and energy-driven transloc ation. ATP depletion inhibits the translocation step. The mechanism of translocation itself and the conformational changes of the NPC compon ents that occur during macromolecular transport, are still unclear. Th e present study investigates the effect of ATP on nuclear pore conform ation in isolated nuclear envelopes from Xenopus laevis oocytes using the atomic force microscope. All experiments were conducted in a salin e solution mimicking the cytosol using unfixed nuclear envelopes. ATP (1 mM) was added during the scanning procedure and the resultant confo rmational changes of the NPCs were directly monitored. Images of the s ame nuclear pores recorded before and during ATP exposure revealed dra matic conformational changes of NPCs subsequent to the addition of ATP . The height of the pores protruding from the cytoplasmic surface of t he nuclear envelope visibly increased while the diameter of the pore o pening decreased. The observed changes occurred within minutes and wer e transient. The slow-hydrolyzing ATP analogue, ATP-gamma-S, in equimo lar concentrations did not exert any effects. The ATP-induced shape ch ange could represent a nuclear pore ''contraction.''.