LANA and hnRNP A1 Regulate the Translation of LANA mRNA through G-Quadruplexes

Abstract

During the latent phase, Kaposi’s sarcoma-associated herpes virus (KSHV) maintains itself inside the host by escaping the host immune surveillance mechanism through restricted protein expression. Latency-associated nuclear antigen (LANA), the most abundantly expressed protein, is essential for viral persistence, as it plays important roles in latent viral DNA replication and efficient segregation of the viral genome to the daughter cells following cell division. KSHV evades immune detection by maintaining the levels of LANA protein below a threshold required for detection by the host immune system but sufficient to maintain the viral genome. LANA achieves this by controlling its expression through regulation of its promoters and by inhibiting its presentation through interaction with the proteins of class I and class II major histocompatibility complex (MHC) pathways. In this study, we identified a mechanism of LANA expression and restricted immune recognition through formation of G-quadruplexes in LANA mRNA. We show that the formation of these stable structures in LANA mRNA inhibits its translation to control antigen presentation, which was supported by treatment of cells with TMPyP4, a G-quadruplex-stabilizing ligand. We identified heterogenous ribonucleoprotein A1 (hnRNP A1) as a G-quadruplex-unwinding helicase, which unfolds these stable secondary structures to regulate LANA translation. IMPORTANCE LANA, the most abundantly expressed protein during latency, is a multifunctional protein which is absolutely required for the persistence of KSHV in the host cell. Even though the functions of LANA in aiding pathogenesis of the virus have been extensively studied, the mechanism of how LANA escapes host’s immune surveillance is not fully understood. This study sheds light on the autoregulatory role of LANA to modulate its expression and immune evasion through formation of G-quadruplexes in its mRNA. We used G-quadruplex-stabilizing ligand to define the inhibition in LANA expression and presentation on the cell surface through MHC class I. We defined the autoregulatory role of LANA and identified a cellular RNA helicase, hnRNP A1, regulating the translation of LANA mRNA. This interaction of hnRNP A1 with LANA mRNA could be exploited for controlling KSHV latency.

Viral G-quadruplexes: New frontiers in virus pathogenesis and antiviral therapy

“…Interestingly, reduced translation has been found to be strictly related to reduction of antigen presentation, one of the main mechanisms adopted by LANA1 to promote immune evasion. 35 Altogether, these data confirm that G4s are key regulatory element in the regulation of host-pathogen interaction in the Herpesviridae family and support the feasible exploitation of these structures as innovative pharmacological targets in the antiherpetic therapy.…”

G-Quadruplexes in RNA Biology: Recent Advances and Future Directions

“…The first exploits the ability of RG4s in both EBNA1 and LANA mRNAs coding sequences to inhibit the translation of these proteins. The structure-immune function relationship was demonstrated by modulating RG4s folding, which then resulted in the altered expression of the viral proteins and, in turn, in modified antigen presentation [85][86][87]. Although the molecular mechanism awaits further investigation, host cell protein factors (hnRNP A1 and nucleolin for LANA and EBNA1 mRNAs, respectively) take part in this regulation by interacting with RG4 structures [86,87].…”

“…The structure-immune function relationship was demonstrated by modulating RG4s folding, which then resulted in the altered expression of the viral proteins and, in turn, in modified antigen presentation [85][86][87]. Although the molecular mechanism awaits further investigation, host cell protein factors (hnRNP A1 and nucleolin for LANA and EBNA1 mRNAs, respectively) take part in this regulation by interacting with RG4 structures [86,87]. This is consistent with the observation that terms associated with viral infection are over-represented in cellular RG4-protein interaction data sets [5,18,19,87].…”

Stealing the Show: KSHV Hijacks Host RNA Regulatory Pathways to Promote Infection

“…G4s have gained some notice for their control over DNA/RNA metabolism. DsDNA viruses have been found to be particularly enriched with putative G4-forming sequences (PQS) [51], and KSHV LANA gene was found to form G4s within its own mRNA [52]. It was demonstrated that LANA protein itself binds to the G4 regions of its own mRNA to prohibit its export to the cytoplasm and thus help control its expression.…”

“…LANA has been shown to modulate antigen presentation by interacting with components of the antigen presentation [66,67]. G4s are now thought to be another way KSHV dampens immune responses by reducing translation of LANA1 and, therefore, this reduces antigen presentation [52]. Together it can be seen that G4s are being used as a regulatory element between host and virus.…”