While performing a yeast two-hybrid library screen to uncover novel MA-interacting proteins, we discovered a specific interaction between a member of the importin beta/karyopherin beta superfamily, importin 9, and the A subunit of PP2A (PR65). This interaction between importin beta and the A subunit was confirmed by in vitro pulldown, immunoprecipitation, and microcystin-Sepharose chromatography. We also found that another family member, importin P, interacted specifically with the A subunit of PP2A. Finally, we showed that treatment of cells with a concentration of okadaic acid known to inhibit PP2A impeded the nuclear localization of an NLS-containing protein. These results provide evidence that these importins can exist in a native complex with endogenous PP2A and that this serine/threonine phosphatase plays a role in regulating the nuclear import of NLS-containing proteins in vivo. (C) 2003 Elsevier Science (USA). All rights reserved.