Novel nuclear import of Vpr promoted by importin alpha is crucial for human immunodeficiency virus type 1 replication in macrophages.
Monocytes/macrophages are major targets of human immunodeficiency virus type 1 (HIV-1) infection. The viral preintegration complex (PIC) of HIV-1 enters the nuclei of monocyte-derived macrophages, but very little PIC migrates into the nuclei of immature monocytes. Vpr, one of the accessory gene products of HIV-1, is essential for the nuclear import of PIC in these cells, although the role of Vpr in the entry mechanism of PIC remains to be clarified. We have shown previously that Vpr is targeted to the nuclear envelope and then transported into the nucleus by importin alpha alone, in an importin beta-independent manner. Here we demonstrate that the nuclear import of Vpr is strongly promoted by the addition of cytoplasmic extract from macrophages but not of that from monocytes and that the nuclear import activity is lost with immunodepletion of importin alpha from the cytoplasmic extract. Immunoblot analysis and real-time PCR demonstrate that immature monocytes express importin alpha at low levels, whereas the expression of three major importin alpha isoforms markedly increases upon their differentiation into macrophages, indicating that the expression of importin alpha is required for nuclear import of Vpr. Furthermore, interaction between importin alpha and the N-terminal alpha-helical domain of Vpr is indispensable, not only for the nuclear import of Vpr but also for HIV-1 replication in macrophages. This study suggests the possibility that the binding of Vpr to importin alpha, preceding a novel nuclear import process, is a potential target for therapeutic intervention.
Nitahara-Kasahara
Yuko
Y
Retrovirus Research Unit, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Kamata
Masakazu
M
Yamamoto
Takuya
T
Zhang
Xianfeng
X
Miyamoto
Yoichi
Y
Muneta
Koho
K
Iijima
Sayuki
S
Yoneda
Yoshihiro
Y
Tsunetsugu-Yokota
Yasuko
Y
Aida
Yoko
Y
D021581:Active Transport, Cell Nucleus; D000818:Animals; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D015870:Gene Expression; D016336:Gene Products, vpr / Q000378:metabolism*; D015497:HIV-1 / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D008264:Macrophages / Q000821:virology*; D018613:Microscopy, Confocal; D011485:Protein Binding; D020033:Protein Isoforms / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014779:Virus Replication; D028901:alpha Karyopherins / Q000096:biosynthesis / Q000378:metabolism*; D054325:vpr Gene Products, Human Immunodeficiency Virus
D016336:Gene Products, vpr; D020033:Protein Isoforms; D012333:RNA, Messenger; D028901:alpha Karyopherins; D054325:vpr Gene Products, Human Immunodeficiency Virus