Inhibition of Polo-like kinase 1 (PLK1) facilitates the elimination of HIV-1 viral reservoirs in CD4 ⁺ T cells ex vivo

Abstract

Although combination antiretroviral therapy is effective in controlling HIV-1 infection, latent HIV-1 proviruses cannot be eliminated. HIV-1 reactivation induced by the mere use of latency-reversing agents is insufficient to render death of reservoir cells, indicating that certain intrinsic survival mechanisms exist. We report that Polo-like kinase 1 (PLK1) plays a critical role in survival of CD4+ T cells that undergo HIV-1 reactivation from latency or de novo infection. PLK1 is elevated in both scenarios, which requires HIV-1 Nef. HIV-1 enhances PLK1 SUMOylation, causing its nuclear translocation and protein stabilization. Inhibition or knockdown of PLK1 markedly facilitates death of HIV-1-infected CD4+ T cells. Furthermore, PLK1 inhibitors strikingly reduce the size of HIV-1 latent reservoirs in primary CD4+ T cells. Our findings demonstrate that HIV-1 infection hijacks PLK1 to prevent cell death induced by viral cytopathic effects, and that PLK1 is a promising target for chemical “killing” of HIV-1 reservoir cells.

Inhibition of polo-like kinase 1 (PLK1) facilitates reactivation of gamma-herpesviruses in B-cell lymphomas and their elimination

“…These cells were treated with doxycycline (2μg/ml) to induce KSHV reactivation. For shRNA assays, endogenous PLK1 was knocked down in AKATA/Bx cells by using its shRNAs expressed from the pINDUCER10 Dox-inducible lentiviral vector, according to the reported protocol 27,14 . Briefly, shRNAs targeting PLK1 (5′-GTT CTT TAC TTC TGG CTA TAT-3′) and the control shRNA targeting firefly luciferase (shNT: 5′-CAC AAA CGC TCT CAT CGA CAA G-3′) were transiently transfected in AKATA/bx cells through electroporation and incubated for 72 hour.…”

“…Other set of B-cells were washed and fixed with 4% paraformaldehyde at RT for 20 min. Pelleted cells were washed and permeabilized with saponin-containing 1× Perm/Wash buffer (BD Biosciences) as described 14 . Cells were then incubated with anti-PLK1 antibody (200μg/ml) diluted in 1× Perm/Wash buffer overnight at 4°C, followed by the incubation with fluorophore-conjugated secondary antibodies for 1 hour at RT in the dark.…”

“…The cytotoxicity of compounds was determined by using the ATP-based CellTiter-Glo Luminescent Cell Viability Assay (Promega) and analyzed by the Cytation 5 multimode reader (luminescent mode). Death of virus-infected or compound-treated cells was determined by using the LIVE/DEAD Fixable Far Red Dead Cell Stain Kit (Invitrogen) and analyzed by the BD Accuri C6 Plus (flow cytometry) 14 .…”

“…Immunoblotting was carried out using the existing protocol 14,25 . In brief, cell pellets were homogenized in ice with 1X RIPA containing protease inhibitor cocktail.…”

“…Polo-like kinase 1 (PLK1) plays an important role especially in cell survival by stabilizing MYC protein in B-cell lymphoma 12 and in tumor cells 13 . Recently we have reported that PLK1 can be elevated during HIV-1 reactivation from latency or de novo infection 14 , which plays a critical role in survival of HIV-infected CD4 + T cells. Although it has been reported that PLK1 expression is also elevated in various human cancers, and abnormally elevated expression of PLK1 is often linked to tumor aggressiveness and poor clinical prognosis, there are scanty information about its role in supporting KSHV or EBV associated cancer progression.…”