Human Immunodeficiency Virus Type 1 Vpr Induces DNA Replication Stress InVitro and In Vivo

Abstract

The human immunodeficiency virus type 1 (HIV-1) viral protein R (Vpr) causes cell cycle arrest in G 2 . Vpr-expressing cells display the hallmarks of certain forms of DNA damage, specifically activation of the ataxia telangiectasia mutated and Rad3-related kinase, ATR. However, evidence that Vpr function is relevant in vivo or in the context of viral infection is still lacking. In the present study, we demonstrate that HIV-1 infection of primary, human CD4؉ lymphocytes causes G 2 arrest in a Vpr-dependent manner and that this response requires ATR, as shown by RNA interference. The event leading to ATR activation in CD4؉ lymphocytes is the accumulation of replication protein A in nuclear foci, an indication that Vpr likely induces stalling of replication forks. Primary macrophages are refractory to ATR activation by Vpr, a finding that is consistent with the lack of detectable ATR, Rad17, and Chk1 protein expression in these nondividing cells. These observations begin to explain the remarkable resilience of macrophages to HIV-1-induced cytopathicity. To study the in vivo consequences of Vpr function, we isolated CD4؉ lymphocytes from HIV-1-infected individuals and interrogated the cell cycle status of anti-p24Gag -immunoreactive cells. We report that infected cells in vivo display an aberrant cell cycle profile whereby a majority of cells have a 4N DNA content, consistent with the onset of G 2 arrest.

HIV-1 Vpr: Mechanisms of G2 arrest and apoptosis

“…Recently, Zimmerman et al reported that HIV-1 infection in primary human CD4+ lymphocytes results in ATR-dependent G 2 arrest and accumulation of RPA, which suggests the presence of stalled replication forks (Zimmerman et al, 2006). Consistent with a model in which Vpr activates the ATR DNA damage response pathway, Yuan et al found that Vpr activates the negative regulator of Cdk1, Wee1 (Yuan et al, 2003).…”

“…Recently, Sakai et al reported that deletion of Vpr, in addition to Vif, rendered HIV-1 incapable of causing G 2 arrest and eliminated the cytopathic properties of the virus, which suggests a link between viral cytopathicity and cell cycle arrest (Sakai et al, 2006). Recently, infected lymphocytes (identified by expression of Gag p24) isolated from recent HIV-1 seroconverters and stained with propidium iodide for DNA content were found to be arrested in G 2 , indicating that G 2 arrest is not merely an in vitro phenomenon (Zimmerman et al, 2006).…”

“…However, the dispensability of ATM in Vpr-induced G 2 arrest (Bartz et al, 1996;Zimmerman et al, 2004), and the lack of ATM activation/phosphorylation in Vpr-expressing G 2 -arrested cells (Lai et al, 2005) strongly support a model in which Vpr activates the checkpoint by a mechanism that does not involve DSBs. In addition, the presence of RPA foci and the strict requirement for ATR and its downstream targets (Lai et al, 2005;Zimmerman et al, 2006), raises the possibility that Vpr, either directly through interaction with chromatin or replication machinery, or indirectly via interference with nuclear structure or the proteosome pathway, causes replication stress.…”

Cellular DNA repair cofactors affecting hepatitis B virus infection and replication

“…Several viruses, including DNA virus herpes simplex v i r u s ( H S V ) [ 1 4 , 1 5 ] , K a p o s i 's s a r c o m a -a s s o c i a t e d herpesvirus [16] , human cytomegalovirus (HCMV) [17] Epstein-Barr virus (EBV) [18] , papillomavirus [19,20] , simian virus 40 (SV40) [21] and retrovirus HIV [22][23][24] , trigger cellular signaling cascades that are characteristic of a DNA damage response. Infection and replication of HBV have been achieved in human hepatoma cell lines and primary fetal human hepatocytes using transfected HBV genomes or HBV serum.…”

Ataxia telangiectasia-mutated-Rad3-related DNA damage checkpoint signaling pathway triggered by hepatitis B virus infection

“…For example, Epstein-Barr Virus (EBV) [16] abrogates the p53 checkpoint signaling pathway through the interaction of the BZLF1 protein and p53 to avoid apoptosis. Other vir uses such as human immunodeficiency virus type 1 (HIV-1) [17][18][19][20] , herpes simplex virus type 1 (HSV-1) [21,22] and human cytomegalovir us [23] can activate and exploit a cellular DNA damage response, which aids viral replication. Adenovirus blocks ATM signaling and concatemer formation through targeting the DNA repair complex of MRN for degradation and mislocalization [24,25] .…”