Cell cycle G2/M arrest through an S phase-dependent mechanism by HIV-1 viral protein R.
BACKGROUND
Cell cycle G2 arrest induced by HIV-1 Vpr is thought to benefit viral proliferation by providing an optimized cellular environment for viral replication and by skipping host immune responses. Even though Vpr-induced G2 arrest has been studied extensively, how Vpr triggers G2 arrest remains elusive.


RESULTS
To examine this initiation event, we measured the Vpr effect over a single cell cycle. We found that even though Vpr stops the cell cycle at the G2/M phase, but the initiation event actually occurs in the S phase of the cell cycle. Specifically, Vpr triggers activation of Chk1 through Ser345 phosphorylation in an S phase-dependent manner. The S phase-dependent requirement of Chk1-Ser345 phosphorylation by Vpr was confirmed by siRNA gene silencing and site-directed mutagenesis. Moreover, downregulation of DNA replication licensing factors Cdt1 by siRNA significantly reduced Vpr-induced Chk1-Ser345 phosphorylation and G2 arrest. Even though hydroxyurea (HU) and ultraviolet light (UV) also induce Chk1-Ser345 phosphorylation in S phase under the same conditions, neither HU nor UV-treated cells were able to pass through S phase, whereas vpr-expressing cells completed S phase and stopped at the G2/M boundary. Furthermore, unlike HU/UV, Vpr promotes Chk1- and proteasome-mediated protein degradations of Cdc25B/C for G2 induction; in contrast, Vpr had little or no effect on Cdc25A protein degradation normally mediated by HU/UV.


CONCLUSIONS
These data suggest that Vpr induces cell cycle G2 arrest through a unique molecular mechanism that regulates host cell cycle regulation in an S-phase dependent fashion.
Li
Ge
G
Department of Pathology, Institute of Human Virology, University of Maryland School of Medicine, Baltimore, MD, USA.
Park
Hyeon U
HU
Liang
Dong
D
Zhao
Richard Y
RY
D018797:Cell Cycle Proteins / Q000378:metabolism; D002455:Cell Division; D000071877:Checkpoint Kinase 1; D016195:G2 Phase; D020868:Gene Silencing; D015497:HIV-1 / Q000472:pathogenicity*; D006367:HeLa Cells; D006801:Humans; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D016196:S Phase; D037521:Virulence Factors / Q000502:physiology*; D020687:cdc25 Phosphatases / Q000378:metabolism; D054325:vpr Gene Products, Human Immunodeficiency Virus / Q000502:physiology*
C467199:CDT1 protein, human; D018797:Cell Cycle Proteins; D037521:Virulence Factors; D054325:vpr Gene Products, Human Immunodeficiency Virus; C514912:vpr protein, Human immunodeficiency virus 1; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; C517004:CDC25A protein, human; C517008:CDC25B protein, human; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases