Elongin C Contributes to RNA Polymerase II Degradation by the Interferon Antagonist NSs of La Crosse Orthobunyavirus

Abstract

Mosquito-borne La Crosse virus (LACV; genus Orthobunyavirus, family Peribunyaviridae, order Bunyavirales) causes up to 100 annual cases of severe meningoencephalitis in children and young adults in the United States. A major virulence factor of LACV is the nonstructural protein NSs, which inhibits host cell mRNA synthesis to prevent the induction of antiviral type I interferons (IFN-α/β). To achieve this host transcriptional shutoff, LACV NSs drives the proteasomal degradation of RPB1, the large subunit of mammalian RNA polymerase II. Here, we show that NSs acts in a surprisingly rapid manner, as RPB1 degradation was commencing already at 1 h postinfection. The RPB1 degradation was partially dependent on the cellular E3 ubiquitin ligase subunit Elongin C. Consequently, removal of Elongin C, but also of the subunits Elongin A or B by siRNA transfection partially rescued general RNAP II transcription and IFN-beta mRNA synthesis from the blockade by NSs. In line with these results, LACV NSs was found to trigger the redistribution of Elongin C out of nucleolar speckles, which, however, is an epiphenomenon rather than part of the NSs mechanism. Our study also shows that the molecular phenotype of LACV NSs is different from RNA polymerase II inhibitors like α-amanitin or Rift Valley fever virus NSs, indicating that LACV is unique in involving the Elongin complex to shut off host transcription and IFN response. IMPORTANCE The mosquito-borne La Crosse virus (LACV; genus Orthobunyavirus, family Peribunyaviridae, order Bunyavirales) is prevalent in the United States and can cause severe childhood meningoencephalitis. Its main virulence factor, the nonstructural protein NSs, is a strong inhibitor of the antiviral type I interferon (IFN) system. NSs acts by imposing a global host mRNA synthesis shutoff, mediated by NSs-driven proteasomal degradation of the RPB1 subunit of RNA polymerase II. Here, we show that RPB1 degradation commences as early as 1 h postinfection, and identify the E3 ubiquitin ligase subunit Elongin C (and its binding partners Elongins A and B) as an NSs cofactor involved in RPB1 degradation and in suppression of global as well as IFN-related mRNA synthesis.

Targeted Mutations in the Fusion Peptide Region of La Crosse Virus Attenuate Neuroinvasion and Confer Protection against Encephalitis

“…The three negative-sense, single-stranded RNA genome segments of LACV each have defined roles in virus pathogenesis. The S segment encodes a virulence factor, NSs, that efficiently inhibits the interferon system of infected mammalian hosts; its absence results in reduced virulence in mice [ 12 , 13 , 14 ]. In at least one instance, neurovirulence mapped to the L segment, which encodes the viral polymerase [ 15 ].…”

An Overview of the Infectious Cycle of Bunyaviruses

“…Similar results were also found when studying LACV [ 84 ]; moreover, this study additionally found that NSs served as a virulence factor only in mammalian cells, with no NSs-related inhibitory activity found in mosquito cells. It should also be noted that NSs of BUNV, LACV, and the related Schmallenberg (SBV) virus were all found to localize to the nucleus of infected cells to inhibit transcription initiation by RNA polymerase II and disrupt nucleoli [ 78 , 85 , 86 , 87 ]. The NSs of Rift Valley fever virus (RVFV), which also interferes with IFN induction and RNA polymerase II activity [ 79 , 88 , 89 ], additionally plays a role in inhibiting mRNA transport [ 90 ].…”

An Overview of the Infectious Cycle of Bunyaviruses

“…Similar results were also found when studying LACV [74]; moreover, this study additionally found that NSs served as a virulence factor only in mammalian cells, with no NSs-related inhibitory activity found in mosquito cells. It should also be noted that NSs of BUNV, LACV and the related Schmallenberg (SBV) virus localize to the nucleus of infected cells, to inhibit transcription initiation by RNA polymerase II and disrupt nucleoli [75][76][77][78]. The NSs of Rift Valley fever virus (RVFV), which also interferes with IFN induction and RNA polymerase II activity [79][80][81], additionally plays a role in inhibiting mRNA transport [82].…”