Human T-cell leukemia virus type 1 tax shuttles between functionally discrete subcellular targets

Human T-cell leukemia virus type 1 (HTLV-1) Tax is a nuclear protein with striking pleiotropic functionality,We recently demonstrated that Tax localizes to a multicomponent nuclear speckled structure (Tax speckled structure [TSS]). Here, we examine these structures further and identify a partial overlap of TSS with transcription hot spots. We used a strategy of directed:expression via fusion proteins to determine if these transcription sites are the subtargets within TSS required for Tax function, When fused to human immunodeficiency virus type 1 (HIV-1) Tat, the resulting Tat-Tax fusion protein displayed neither a Tat-like nor a Tax-like pattern but rather was targeted specifically to the transcription subsites, The Tat-Tax fusion was able to activate both the HIV-1 long terminal repeat (LTR) and the HTVL-1 LTR at the same level as the individual component; thus, targeting proteins to transcription hot spots was compatible with both Tax and Tat transcription function. In contrast, the fusion with HIV-1 Rev, Rev Tax,: resulted in a pattern of expression that was largely Rev-like (nucleolar and cytoplasmic), The reduced localization of Rev-Tax to transcription sites was reflected in a 10-fold drop in activation of the HTLV-1LTR. However, there was no loss in the ability of Tax to activate via NF-kappa B, Thus, NF-kappa B-dependent Tax function does not require targeting of Tax to these transcription sites and suggests that activation via NF-kappa B is a cytoplasmic function. Selective mutation of the nuclear localization signal site in the Rev portion resulted in retargeting of Rev-Tax to TSS and subsequent restoration of transcription function, demonstrating that inappropriate localization preceded loss of function. Mutation of the nuclear export signal site in the Rev-portion had no effect on transcription, although the relative amount of Rev-Tax in the cytoplasm was reduced. Finally,:in explaining how Tax can occupy multiple subcellular sites, we show that Tax shuttles from the nucleus to the cytoplasm in a heterokaryon fusion assay. Thus, pleiotropic functionality by Tax is regulatable via shuttling between discrete subcellular compartments.