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Nuclear protein import is dependent on specific targeting signals within cargo proteins recognized by importins (IMPs) that mediate translocation through the nuclear pore. Recent evidence, however, implicates a role for the microtubule (MT) network in facilitating nuclear import of the cancer regulatory proteins parathyroid hormone‐related protein (PTHrP) and p53 tumor suppressor. Here we assess the extent to which MT and actin integrity may be generally required for nuclear protein import for the first time. We examine 10 nuclear‐localizing proteins with diverse IMP‐dependent nuclear import pathways, our results indicating that the cytoskeleton does not have a general mechanistic role in nuclear localization sequence‐dependent nuclear protein import. Of the proteins examined, only the p110_Rb tumor suppressor protein Rb, together with p53 and PTHrP, was found to require MT integrity for optimal nuclear …