IFN-independent G0 arrest and SAMHD1 activation following TLR4 activation in macrophages

Monocyte-derived macrophages mostly reside in a resting, G0 state, expressing high levels of dephosphorylated, active SAMHD1. We have previously shown that macrophages can re-enter the cell cycle without division, into a G1-like state. This cell cycle re-entry is accompanied by phosphorylation of the dNTP hydrolase/ antiviral restriction factor SAMHD1 at T592 by the cyclin-dependent kinase CDK1. HIV-1 successfully infects macrophages in G1 through exploiting this naturally occurring window of opportunity where SAMHD1 antiviral activity is de-activated. Here we demonstrate for the first time that LPS activation of the pathogen associated molecular pattern (PAMP) receptor TLR4 induces G0 arrest in human macrophages. We show this G0 arrest is MyD88-independent and therefore NFkB independent. Furthermore, the effect of TLR4 activation on cell cycle is regulated by (a) the canonical IFN-dependent pathway following TBK1 activation and IRF3 translocation and (b) an IFN-independent pathway that occurs prior to TBK1 activation, and that is accompanied by CDK1 downregulation, p21 upregulation and SAMHD1 dephosphorylation at T592. Furthermore, we show by siRNA knockdown of SAMHD1 that the interferon independent pathway activated by TLR4 is able to potently block HIV-1 infection in macrophages specifically via SAMHD1. Finally, ingestion of whole E. Coli and TLR4 activation by macrophages also activates SAMHD1 via the interferon independent pathway. Together, these data demonstrate that macrophages can rapidly activate an intrinsic cell arrest and anti-viral state by activation of TLR4 prior to IFN secretion, thereby highlighting the importance of cell cycle regulation as a response to danger signals in human macrophages. Interferon independent activation of SAMHD1 by TLR4 represents a novel mechanism for limiting the HIV-1 reservoir size and should be considered for host-directed therapeutic approaches that may contribute to curative interventions.