HUMAN FOAMY VIRUS BELL TRANSACTIVATOR CONTAINS A BIPARTITE NUCLEAR-LOCALIZATION DETERMINANT WHICH IS SENSITIVE TO PROTEIN CONTEXT AND TRIPLE MULTIMERIZATION DOMAINS

The Bel1 protein of human foamy virus is a 300-amino-acid nuclear regu latory protein which transactivates the gene expression directed by th e homologous long terminal repeat and the human immunodeficiency virus type 1 long terminal repeat. While previous reports suggested that th e single basic domain of Bell from residues 211 to 222 and/or 209 to 2 26 is necessary and sufficient for efficient nuclear localization (L. K. Venkatesh, C. Yang, P. A. Theodorakis, and G. Chinnandurai, J. Viro l. 67:161-169, 1993; P. He, J. D. Sun, E. D. Garrett, and B. R. Cullen , J. Virol. 67:1896-1904, 1993), our recent data showed that another b asic domain, from amino acid residues 199 to 200, is also required for nuclear localization of Bell (C. W. Lee, C. Jun, K. J. Lee, and Y. C. Sung, J. Virol. 68:2708-2719, 1994). To clarify this discrepancy, we constructed various bell-lacZ chimeric constructs and several linker i nsertion mutants and determined their subcellular localization. When t he region of Bell containing basic domains was placed at an internal s ite of the lacZ gene, the nuclear localization signal (NLS) of Bell co nsisted of two discontinuous basic regions separated by an intervening sequence. Moreover, insertion of specific amino acids between two bas ic regions disrupted the activity of the Bell NLS. On the other hand, Bell residues 199 and 200 were not required to direct the Bell-beta-ga lactosidase chimeric protein to the nucleus when the Bell NLS was appe nded to the amino terminus of beta-galactosidase. These, results indic ate that the function of the Bell NLS is sensitive to the protein cont ext within which the sequence is present. In addition, we demonstrated that the Bell protein forms a multimeric complex in the nuclei of mam malian cells by using a sensitive in vivo protein-protein interaction assay. Mutational analyses revealed that the regions which mediate mul timer formation map to three domains of Bell, i.e., residues 1 to 31, 42 to 82, and 82 to 111. Furthermore, our results show that the region of Bell from residues 202 to 226 prevents Bell from forming a multime ric complex.