IDENTIFICATION OF THE NUCLEAR-LOCALIZATION SIGNAL OF THE POU DOMAIN PROTEIN TST-1 OCT6/

POU domain proteins are important regulators of development and termin al differentiation based upon their transcriptional activity in the nu cleus. Here, we analyzed the mechanism underlying the nuclear localiza tion of Tst-1/Oct6, a member of this family that regulates events duri ng neurogenesis and myelination. Nuclear localization of Tst-1/Oct6 wa s dependent on the POU domain, as its deletion prevented access to the nucleus, whereas its transfer to the amino terminus of beta-galactosi dase was sufficient to prompt nuclear accumulation of this normally cy tosolic protein. Interestingly, nuclear localization and high affinity DNA binding were two independent functions of the POU domain and coul d be separated in several mutants. While specific high affinity bindin g to DNA required the presence of both the POU-specific and the POU ho meodomain, the POU-specific domain was dispensable for nuclear localiz ation of Tst-1/Oct6. Rather, the nuclear localization function was sel ectively contained within the POU homeodomain. Specifically, a basic c luster (GRKRKKRT) preceding helix 1 of the homeodomain was shown by de letion mutagenesis to be involved in the nuclear localization of Tst-1 /Oct6. This sequence, which is highly conserved among POU domain prote ins, was by itself capable of translocating beta-galactosidase to the nucleus defining it as the bona fide nuclear localization signal of Ts t-1/Oct6 and presumably other POU domain factors.