ROLE OF HTLV-1 P30 DURING INFECTION OF HUMAN MONOCYTES AND DENDRITIC CELLS

Human T-Lymphotropic virus type 1 (HTLV-1) is the causative agent of two distinct pathologies, adult T-cell leukemia (ATL), an aggressive malignancy of mature CD4+ T-cells, and tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM), a demyelinating neurodegenerative disease. An estimated 10–20 million people worldwide are infected with HTLV-1. While the majority of HTLV-1- infected individuals remain asymptomatic, a low percentage of patients develop either ATL (3–5%) or TSP/HAM (0.3–2%) after a long period of clinical latency. HTLV-1 primarily infects CD4+ T-cells and has been detected in ex vivo CD8+ Tcells, B-cells, monocytes and dendritic cells (DC) from infected individuals. These cells may contribute, more than previously thought, to viral persistence and pathogenesis. DCs isolated from HTLV-1-positive patients are defective in IFN- response upon in vitro stimulation and their ability to activate T-cells is impaired. In addition, in HTLV-1 infected patients alteration in monocytes differentiation and activation has been reported. Contrarily to how observed in T or B cell, where HTLV-1 p30 is not required in vitro for an efficient viral replication, p30 seems to play a key role during HTLV-1 infection of DCs. In the macaque model, the ablation of p30 expression within the HTLV-1 provirus (p30-KO) or p12/p8 (p12-KO) severely affects infectivity. Furthermore, the infectivity of p30-KO in human primary monocyte-derived dendritic cells (Mo-mDCs) is severely impaired and is not sustained over time. Prior data demonstrated that p30, by interacting with the cellular transcription factor PU.1 in human monocytes, affects TLR4 signaling and the expression of several genes involved in apoptosis, cell cycle, and transcription, pointing to p30 as a regulator of innate response to HTLV-1. The interaction of p30 and PU.1 might lead to inhibition of the transcriptional activity of PU.1. We used a human monocytic cell line, THP-1 and primary dendritic cells, to study the mechanism of p30 and p12/p8 requirement in these cell types. We hypothesized that p30 and/or p12/p8 affected not only TLR3/4 but also TLR7/8 signaling in virus infected monocytes and in dendritic cells, and that by affecting the type 1 IFN response p30 and/or p12/p8 affects the activation and differentiation of these cells and ultimately the host response to the virus allowing viral replication and spread throughout the body. We confirm that p30 decreases the expression of TLR4 on THP-1 surface and the production of IL12 and TNF by these cells. p30 is also able to inhibit the expression of interferon responsive genes (ISG) following stimulation with LPS of the TLR4 and with polyI:C of TLR3, but not of TLR7/8 with Imiquimod. Results in THP-1 mirrored those in ex-vivo human primary monocyte-derived dendritic cells (Mo-mDC). The effect of p30 on TLRs signaling was also demonstrated by ablating its expression within a molecular clone of HTLV-1. HTLV-1 infection of monocytes inhibited TLR3 and TLR4–induced ISGs expression by 50 to 90%, depending on the genes, whereas the isogenic clone p30 knockout (KO) virus was less effective in inhibition of TLR3 and TRL4 signaling and displayed lower infectivity. Viral expression and inhibition of ISGs transcription, was however, rescued by restoration of p30 expression. A chromatin immunoprecipitation (ChIP) assay demonstrated that p30 inhibits initiation and elongation of PU.1 dependent transcription of interferon- 1 (IFN 1), IFN , and TLR4 genes upon TLRs stimulation. In contrast, experiments conducted with p12/p8 did not demonstrate an effect on IFN response gene expression. These results provide a mechanistic explanation on the requirement of p30 for HTLV-1 infectivity in vivo, and suggest that dampening interferon responses in monocytes and DCs is specific for p30 and represent an essential early step for permissive HTLV-1 infection and persistence.