Crotonylation Sensitizes IAPI-Induced Disruption of Latent HIV by Modulating ncNF-κB Signaling at the Step of p100 Cleavage Into p52

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The eradication of infection with human immunodeficiency virus type 1 (HIV) is difficult to achieve, largely due to stable viral reservoirs. While many strategies have been tested, a clinical meaningful success has yet to be achieved, indicating that our understanding of HIV latency is incomplete and strategies to disrupt latency are not yet ideal. In this report, we show that crotonylation, a recently discovered epigenetic modification involved in HIV transcription from latency, also acts to enhance AZD5582-induced noncanonical NF-κB (ncNF-κB) signaling, further augmenting HIV latency reversal via this pathway. This was tested in Jurkat and U1 cell line models of latency, HIV latently infected primary CD4+ T cells and resting CD4+ T cells isolated from ART-suppressed people living with HIV ex vivo . Surprisingly, crotonylation upregulates the levels of the active p52 subunit of the NF-κB transcription factor induced by AZD5582 in Jurkat models of HIV latency. Quantitative mass spectrometry screening and biochemical analysis suggest that the ubiquitination E3 ligase TRIM27 may be involved in p100 cleavage into p52 activated by AZD5582, part of the heterodimer to activate NF-κB for the transcription of HIV. When TRIM27 is depleted, transcription of HIV and AZD5582-induced latency reversal are reduced. Interestingly, both p100 and p52 levels are reduced by TRIM27 siRNA knockdown or by pharmacologic inhibition of TRIM27 deubiquitinase partner protein USP7 during the induction of ncNF-κB signaling. These observations reveal the complexity of HIV transcriptional machinery, particularly of NF-κB factors known to be essential for HIV latency disruption.