Morozov, V. | Search bioRxiv

bioRxiv2026

Non-canonical histone H3.3 and its chaperones HIRA and DAXX participate in the regulation of KSHV latency

Kaposis sarcoma-associated herpesvirus (KSHV), also named HHV-8, is the etiological agent of Kaposi sarcoma (KS), Primary effusion lymphoma (PEL), and Multicentric Castlemans disease. After de novo infection, KSHV genomes rapidly circularize and acquire a chromatin state that favors latency. During latency, the KSHV episome is decorated with distinct epigenetic marks that segregate the viral genome into transcriptionally active and repressed domains, enabling persistent silencing of lytic genes while retaining the capacity for reactivation. Transcription activity of chromatin is regulated at multiple levels, including the incorporation of histone variants such as H3.3, by a specific set of histone chaperones such as HIRA and DAXX. The interaction between LANA and these interphase active chaperones suggests that H3.3 deposition is a critical driver of early chromatinization and the long-term stability of KSHV latency. We detected rapid H3.3 deposition on KSHV episomes and on episomes within long-term infected cells. Moreover, we demonstrated that genetically disrupting the host H3.3 chaperone HIRA pathway by CRISPR/Cas9-mediated knockout impacted the regulation of LANA and maintenance of viral latency that was not altered in DAXX knockout cells. Collectively, these results support a role for HIRA-mediated H3.3 deposition in the regulation of KSHV latency.

molecular biology↗

bioRxiv2018

Human Norovirus Neutralized by a Monoclonal Antibody Targeting the HBGA Pocket

Temporal changes in the GII.4 human norovirus capsid sequences occasionally result in the emergence of genetic variants capable of causing new epidemics. The GII.4 persistence is believed to be associated with the recognition of numerous histo-blood group antigen (HBGA) types and antigenic drift. We found that one of the earliest known GII.4 isolate (1974) and a more recent epidemic GII.4 variant (2012) had varied norovirus-specific monoclonal antibody (MAb) reactivities, yet similar HBGA binding profiles. To better understand the binding interaction of one MAb (10E9) that had varied reactivities with these GII.4 variants, we determined the X-ray crystal structure of the NSW-2012 GII.4 P domain 10E9 Fab complex. We showed that the 10E9 Fab interacted with conserved and variable residues, which could be associated with antigenic drift. Interestingly, the 10E9 Fab binding pocket partially overlapped the HBGA pocket and had direct competition for conserved HBGA binding residues (i.e., Arg345 and Tyr444). Indeed, the 10E9 MAb blocked norovirus VLPs from binding to several sources of HBGAs. Moreover, the 10E9 antibody completely abolished virus replication in the human norovirus intestinal enteroid cell culture system. Our new findings provide first direct evidence that competition for GII.4 HBGA binding residues and steric obstruction could lead to norovirus neutralization. On the other hand, the 10E9 MAb recognized residues flanking the HBGA pocket, which are often substituted as the virus evolves. This mechanism of antigenic drift likely influences herd immunity and impedes the possibility of acquiring broadly reactive HBGA-blocking antibodies.\n\nIMPORTANCEThe emergence of new epidemic GII.4 variants is thought to be associated with changes in antigenicity and HBGA binding capacity. Here, we show that HBGA binding profiles remain unchanged between 1974 and 2012 GII.4 variants, whereas these variants showed varying levels of reactivities against a panel of GII.4 MAbs. We identified a MAb that bound at the HBGA pocket and blocked norovirus VLPs from binding to HBGAs and neutralized norovirus virions in the cell culture system. Raised against GII.4 2006 strain this MAb was unreactive to GII.4 1987 isolate, but was able to neutralize newer 2012 strain, which has important implications for vaccine design. Altogether, these new findings suggested that the amino acid variations surrounding HBGA pocket lead to temporal changes in antigenicity without affecting the ability of GII.4 variants to bind HBGAs, which are known co-factors for infection.

molecular biology↗