Identification of nuclear-import and cell-cycle regulatory proteins that bind to prothymosin alpha

Prothymosin alpha (ProT alpha) is a nuclear protein that is widely distribu ted in mammalian tissues, and is thought to play a role in cell proliferati on. In an attempt to shed light on this role, affinity chromatography on Pr oT alpha -Sepharose columns was used to identify proteins in subcellular ex tracts of transformed human lymphocytes (NC37 cells) that interact with Pro T alpha in vitro, and thus may interact with ProT alpha in vivo. Immunoblot ting techniques were used to screen the ProT alpha -binding fractions for h istones and other proteins involved in nuclear transport and cell-cycle con trol. The most abundant ProT alpha-binding proteins were histones H2A, H2B, H3, and H4. Of the nuclear-transport proteins, karyopherin beta (1), Rch-1 , Ran, and RCC1 were detected at high concentrations; NTF2, nucleoporin p62 , and Hsp70 were detected at low concentrations; while tranportin, CAS, and Ran BP1 were not detected. Of the cell-cycle control proteins, PCNA, Cdk2, and cyclin A were detected at high concentrations; cdc2, Cdk4, and cyclin B were detected at very low concentrations; while cyclin D1, cyclin D3, Cip 1, and Kip1 were not detected. These results suggest (i) that ProT alpha is transported into the nucleus by the karyopherin beta (1) - Rch-1 complex, and (ii) that ProT alpha may interact in the nucleus with proteins involved in DNA metabolism and cell-cycle control.