2012. spite of decreased SHP1 levels in gammaherpesvirus-driven B Incyclinide cell lymphomas, B cell-intrinsic SHP1 expression plays a proviral role during the establishment of chronic infection, suggesting that the gammaherpesvirus-SHP1 interaction is more nuanced and is modified by the stage of infection and pathogenesis. IMPORTANCE Gammaherpesviruses establish lifelong infection in a majority of adults worldwide and are associated with a number Incyclinide of malignancies, including B cell lymphomas. These viruses infect naive B cells and manipulate B cell differentiation to achieve a lifelong infection of memory B cells. The germinal center stage of B cell differentiation is important as both an amplifier of the viral latent reservoir and the target of malignant transformation. In this study, we demonstrate that expression of tyrosine phosphatase SHP1, a negative regulator that normally limits the activation and proliferation of hematopoietic cells, enhances the gammaherpesvirus-driven germinal center response and the establishment of chronic infection. The results of this study uncover an intriguing beneficial interaction between gammaherpesviruses that are presumed to profit from B cell activation and a cellular KSHV ORF26 antibody phosphatase that is traditionally perceived to be a negative regulator of the same processes. studies of human gammaherpesviruses. Thus, the current study utilizes MHV68, a natural rodent pathogen that is genetically and biologically similar to EBV and KSHV (35,C37). After a brief acute lytic replication in a naive host, MHV68 establishes latency in several organs, including the spleen (38, 39). Viral latency in the spleen peaks at 14 to 18?days postinfection, with most of the latent virus being present in the germinal center B cells (40, 41). To define the role of SHP1 in gammaherpesvirus infection while overcoming the deleterious effects of global SHP1 deficiency, a published mouse model of B cell-specific SHP1 deficiency was used (33). To determine the effect of B cell-specific SHP1 deficiency on the establishment of MHV68 latency, SHP1flox/flox (SHP1fl/fl) mice heterozygous for CD19 promoter-driven Cre recombinase or homozygous for wild-type (wt) CD19 allele (referred to as CD19 Cre-positive and CD19 Cre-negative mice, respectively, throughout this article) were infected with MHV68, and parameters of viral latency were determined at 16?days postinfection. In spite of the known role of SHP1 as a negative regulator of B cell activation, with the latter supporting the establishment of chronic gammaherpesvirus infection, CD19 Cre-positive mice displayed a significantly lower frequency of MHV68 DNA-positive splenocytes than CD19 Cre-negative mice (11-fold; Fig. 1A), along with a decrease in the absolute number of MHV68 DNA-positive splenocytes (7-fold; Fig. 1B). Similarly, the frequency of reactivation from CD19 Cre-positive splenocytes was decreased compared to that in the control group (Fig. 1C). Thus, B cell-specific SHP1 deficiency resulted in the overall attenuation of MHV68 latency and reactivation. Open in a separate window FIG 1 Loss of SHP1 expression in B cells leads to attenuated establishment of MHV68 chronic infection. CD19 Cre-negative or CD19 Cre-positive mice were intranasally infected with 500 PFU of MHV68, and splenocytes were harvested at 16?days postinfection. As described in Materials and Methods, limiting dilution assays were used to measure the frequency (A) and, subsequently, the absolute number (B) of MHV68 genome-positive splenocytes and the frequency of viral reactivation (C). Splenocytes from 3 to 5 5 mice were pooled within an individual group in each experiment, and data from at least 3 independent experiments were pooled. Error bars here and throughout the figures represent the standard error of the measurement. The Incyclinide dashed lines in panels A and C are drawn at 63% to define the frequency of a positive event. CPE, cytopathic effect. B cell-intrinsic SHP1 expression supports the MHV68-driven germinal center response. Gammaherpesviruses exploit B cell differentiation via latent infection of naive B cells, with the subsequent entry of both infected and uninfected naive B cells into the germinal center response. The rapid proliferation of germinal center B cells passively expands the latent viral reservoir (8), such that the germinal center B cells host a majority of latent MHV68 at 16?days postinfection. Having observed a decreased frequency of MHV68 DNA-positive splenocytes, we next examined the germinal center response. As previously published (33), B cell-specific SHP1 deficiency results in an increase in the splenic B-1 B cell population that expresses intermediate.