HIV reprograms host m6Am RNA methylome by viral Vpr protein-mediated degradation of PCIF1

Abstract

N6,2′-O-dimethyladenosine (m6Am) is an abundant RNA modification located adjacent to the 5′-end of the mRNA 7-methylguanosine (m7G) cap structure. m6A methylation on 2′-O-methylated A at the 5′-ends of mRNAs is catalyzed by the methyltransferase Phosphorylated CTD Interacting Factor 1 (PCIF1). The role of m6Am and the function of PCIF1 in regulating host–pathogens interactions are unknown. Here, we investigate the dynamics and reprogramming of the host m6Am RNA methylome during HIV infection. We show that HIV infection induces a dramatic decrease in m6Am of cellular mRNAs. By using PCIF1 depleted T cells, we identify 2237 m6Am genes and 854 are affected by HIV infection. Strikingly, we find that PCIF1 methyltransferase function restricts HIV replication. Further mechanism studies show that HIV viral protein R (Vpr) interacts with PCIF1 and induces PCIF1 ubiquitination and degradation. Among the m6Am genes, we find that PCIF1 inhibits HIV infection by enhancing a transcription factor ETS1 (ETS Proto-Oncogene 1, transcription factor) stability that binds HIV promoter to regulate viral transcription. Altogether, our study discovers the role of PCIF1 in HIV–host interactions, identifies m6Am modified genes in T cells which are affected by viral infection, and reveals how HIV regulates host RNA epitranscriptomics through PCIF1 degradation.

Anomalous HIV-1 RNA, How Cap-Methylation Segregates Viral Transcripts by Form and Function

“…The cap-specific adenosine methyltransferase phosphorylated CTD-interacting factor 1 (PCIF1) catalyzes methylation of the cap-proximal adenosine to m 7 Gpppm 6 A m on host mRNAs involved in transcriptional regulation and stress response [ 62 , 63 ]. Moreover, PCIF1-m 6 A m methylation has been shown to attenuate interferon-β–mediated suppression of viral infection.…”

“…VSV and RABV mRNAs are substrates of PCIF1, indicating PCIF1 activity contributes to viral evasion of innate immune suppression. PCIF1 methylation activity on host substrates was found to restrict HIV replication [ 63 ]. Analysis of the HIV-1 TSS initially identified guanosine-guanosine as the primary sequence at the 5′ end of HIV-1 in lymphocytes [ 65 , 66 , 67 ].…”

“…Thus, PCIF1 improbably contributes to HIV evasion of innate immune suppression directly [ 69 ]. HIV accessory viral protein R (Vpr) has been shown to counter PCIF1 repression of proviruses by triggering its ubiquitination and degradation through a proteasome pathway [ 63 ]. Genome-wide screens of PCIF1-knockout cells identified several transcription factors, e.g., ETS1 and HUSH family components.…”

“…Vpr also counteracted transcriptional repression of proviruses by the HUSH complex that may also be regulated by PCIF1 [ 69 ]. Experiments are warranted to fully investigate PCIF1 activity on HUSH components and other HIV host dependency and restriction factors [ 63 ].…”

Bimodal Expression Patterns, and Not Viral Burst Sizes, Predict the Effects of Vpr on HIV-1 Proviral Populations in Jurkat Cells

“…Some tissue culture models for HIV-1 latency and reactivation, such as the widely used J-Lat clones, assess long terminal repeat (LTR) promoter activity by reporter gene expression but lack genetic elements believed to be dispensable for HIV-1 latency ( 24 – 26 ). One such element is the vpr gene, whose product plays roles in viral infectivity in vivo ( 27 – 30 ) but also causes cell cycle arrest and can induce widespread changes in host gene expression ( 31 – 33 ). Many latency models use vpr -defective proviruses, which, when cultured over time to allow proviral silencing, can be used for reactivation studies ( 34 – 37 ).…”