The nuclear function of the nuclear diffusion inhibitory signal of human immunodeficiency virus type 1: critical roles in dominant nuclear localization and intracellular stability.

OBJECTIVES: Human immunodeficiency virus type 1 (HIV-1) Rev is a nucleocytoplasmic shuttling protein with dominant localization in the cell nucleus/nucleolus. To clarify the mechanism(s) that enables such a biased subcellular localization under the co-presence of nuclear/nucleolar targeting (NOS) and nuclear export signals (NES), the properties of another functional domain, a nuclear diffusion inhibitory signal (NIS), was investigated. STUDY DESIGN: The NIS was previously shown to interfere with passive nuclear entry of small proteins. Intracellular distribution and degradation profiles of Rev mutants that harbor mutations in the NIS were analyzed biochemically and cellbiologically. RESULTS: A NIS-deficient Rev mutant, which was no longer functional as Rev, displayed a significantly more rapid degradation profile as a consequence of its dramatic leakage into the cytoplasm. Additionally, disabling the NOS/nuclear localization signal (NLS), as well as the NIS, resulted in further rapid degradation, which also supports the hypothetical role of the nucleolus as a Rev storage site. CONCLUSIONS: It was uncovered that the NIS is playing a key role in HIV-1 Rev function by maintaining the nuclear-dominant localization and the intracellular stability of Rev.