Murine Gamma-herpesvirus Immortalization of Fetal Liver-Derived B Cells Requires both the Viral Cyclin D Homolog and Latency-Associated Nuclear Antigen

Human gammaherpesviruses are associated with the development of lymphoproliferative diseases and B cell lymphomas, particularly in immunosuppressed hosts. Understanding the molecular mechanisms by which human gammaherpesviruses cause disease is hampered by the lack of convenient small animal models to study them. However, infection of laboratory strains of mice with the rodent virus murine gammaherpesvirus 68 (MHV68) has been useful in gaining insights into how gammaherpesviruses contribute to the genesis and progression of lymphoproliferative lesions. In this report we make the novel observation that MHV68 infection of murine day 15 fetal liver cells results in their immortalization and differentiation into B plasmablasts that can be propagated indefinitely in vitro, and can establish metastasizing lymphomas in mice lacking normal immune competence. The phenotype of the MHV68 immortalized B cell lines is similar to that observed in lymphomas caused by KSHV and resembles the favored phenotype observed during MHV68 infection in vivo. All established cell lines maintained the MHV68 genome, with limited viral gene expression and little or no detectable virus production - although virus reactivation could be induced upon crosslinking surface Ig. Notably, transcription of the genes encoding the MHV68 viral cyclin D homolog (v-cyclin) and the homolog of the KSHV latency-associated nuclear antigen (LANA), both of which are conserved among characterized γ2-herpesviruses, could consistently be detected in the established B cell lines. Furthermore, we show that the v-cyclin and LANA homologs are required for MHV68 immortalization of murine B cells. In contrast the M2 gene, which is unique to MHV68 and plays a role in latency and virus reactivation in vivo, was dispensable for B cell immortalization. This new model of gammaherpesvirus-driven B cell immortalization and differentiation in a small animal model establishes an experimental system for detailed investigation of the role of gammaherpesvirus gene products and host responses in the genesis and progression of gammaherpesvirus-associated lymphomas, and presents a convenient system to evaluate therapeutic modalities

Global mRNA Degradation during Lytic Gammaherpesvirus Infection Contributes to Establishment of Viral Latency

During a lytic gammaherpesvirus infection, host gene expression is severely restricted by the global degradation and altered 3′ end processing of mRNA. This host shutoff phenotype is orchestrated by the viral SOX protein, yet its functional significance to the viral lifecycle has not been elucidated, in part due to the multifunctional nature of SOX. Using an unbiased mutagenesis screen of the murine gammaherpesvirus 68 (MHV68) SOX homolog, we isolated a single amino acid point mutant that is selectively defective in host shutoff activity. Incorporation of this mutation into MHV68 yielded a virus with significantly reduced capacity for mRNA turnover. Unexpectedly, the MHV68 mutant showed little defect during the acute replication phase in the mouse lung. Instead, the virus exhibited attenuation at later stages of in vivo infections suggestive of defects in both trafficking and latency establishment. Specifically, mice intranasally infected with the host shutoff mutant accumulated to lower levels at 10 days post infection in the lymph nodes, failed to develop splenomegaly, and exhibited reduced viral DNA levels and a lower frequency of latently infected splenocytes. Decreased latency establishment was also observed upon infection via the intraperitoneal route. These results highlight for the first time the importance of global mRNA degradation during a gammaherpesvirus infection and link an exclusively lytic phenomenon with downstream latency establishment

The Scientific Legacy of NASA’s Operation IceBridge

The National Aeronautics and Space Administration (NASA)’s Operation IceBridge (OIB) was a 13-year (2009–2021) airborne mission to survey land and sea ice across the Arctic, Antarctic, and Alaska. Here, we review OIB’s goals, instruments, campaigns, key scientific results, and implications for future investigations of the cryosphere. OIB’s primary goal was to use airborne laser altimetry to bridge the gap in fine-resolution elevation measurements of ice from space between the conclusion of NASA’s Ice, Cloud, and land Elevation Satellite (ICESat; 2003–2009) and its follow-on, ICESat-2 (launched 2018). Additional scientific requirements were intended to contextualize observed elevation changes using a multisensor suite of radar sounders, gravimeters, magnetometers, and cameras. Using 15 different aircraft, OIB conducted 968 science flights, of which 42% were repeat surveys of land ice, 42% were surveys of previously unmapped terrain across the Greenland and Antarctic ice sheets, Arctic ice caps, and Alaskan glaciers, and 16% were surveys of sea ice. The combination of an expansive instrument suite and breadth of surveys enabled numerous fundamental advances in our understanding of the Earth’s cryosphere. For land ice, OIB dramatically improved knowledge of interannual outlet-glacier variability, ice-sheet, and outlet-glacier thicknesses, snowfall rates on ice sheets, fjord and sub-ice-shelf bathymetry, and ice-sheet hydrology. Unanticipated discoveries included a reliable method for constraining the thickness within difficult-to-sound incised troughs beneath ice sheets, the extent of the firn aquifer within the Greenland Ice Sheet, the vulnerability of many Greenland and Antarctic outlet glaciers to ocean-driven melting at their grounding zones, and the dominance of surface-melt-driven mass loss of Alaskan glaciers. For sea ice, OIB significantly advanced our understanding of spatiotemporal variability in sea ice freeboard and its snow cover, especially through combined analysis of fine-resolution altimetry, visible imagery, and snow radar measurements of the overlying snow thickness. Such analyses led to the unanticipated discovery of an interdecadal decrease in snow thickness on Arctic sea ice and numerous opportunities to validate sea ice freeboards from satellite radar altimetry. While many of its data sets have yet to be fully explored, OIB’s scientific legacy has already demonstrated the value of sustained investment in reliable airborne platforms, airborne instrument development, interagency and international collaboration, and open and rapid data access to advance our understanding of Earth’s remote polar regions and their role in the Earth system

Cross-species conservation of episome maintenance provides a basis for in vivo investigation of Kaposi's sarcoma herpesvirus LANA

Many pathogens, including Kaposi’s sarcoma herpesvirus (KSHV), lack tractable small animal models. KSHV persists as a multi-copy, nuclear episome in latently infected cells. KSHV latency-associated nuclear antigen (kLANA) binds viral terminal repeat (kTR) DNA to mediate episome persistence. Model pathogen murine gammaherpesvirus 68 (MHV68) mLANA acts analogously on mTR DNA. kLANA and mLANA differ substantially in size and kTR and mTR show little sequence conservation. Here, we find kLANA and mLANA act reciprocally to mediate episome persistence of TR DNA. Further, kLANA rescued mLANA deficient MHV68, enabling a chimeric virus to establish latent infection in vivo in germinal center B cells. The level of chimeric virus in vivo latency was moderately reduced compared to WT infection, but WT or chimeric MHV68 infected cells had similar viral genome copy numbers as assessed by immunofluorescence of LANA intranuclear dots or qPCR. Thus, despite more than 60 Ma of evolutionary divergence, mLANA and kLANA act reciprocally on TR DNA, and kLANA functionally substitutes for mLANA, allowing kLANA investigation in vivo. Analogous chimeras may allow in vivo investigation of genes of other human pathogens