DETERMINATION OF THE FUNCTIONAL DOMAIN ORGANIZATION OF THE IMPORTIN-ALPHA NUCLEAR IMPORT FACTOR

Although importin alpha (Imp alpha) has been shown to act as the recep tor for basic nuclear localization signals (NLSs) and to mediate their recruitment to the importin beta nuclear import factor, little is kno wn about the functional domains present in Imp alpha, with the excepti on that importin beta binding is known to map close to the Imp alpha N H2 terminus. Here, we demonstrate that sequences essential for binding to the CAS nuclear export factor are located near the Imp alpha COOH terminus and include a critical acidic motif. Although point mutations introduced into this acidic motif inactivated both CAS binding and Im p alpha nuclear export, a putative leucine-rich nuclear export signal proved to be neither necessary nor sufficient for Imp alpha nuclear ex port. Analysis of sequences within Imp alpha that bind to the SV-40 T antigen NLS or to the similar LEF-1 NLS revealed that both NLSs intera ct with a subset of the eight degenerate armadillo (Arm) repeats that form the central part of Imp a. However, these two NLS-binding sites s howed only minimal overlap, thus suggesting that the degeneracy of the Arm repeat region of Imp alpha may serve to facilitate binding to sim ilar but nonidentical basic NLSs. Importantly, the SV-40 TNLS proved a ble to specifically inhibit the interaction of Imp alpha with CAS in v itro, thus explaining why the SV-40 T NLS is unable to also function a s a nuclear export signal.