LANA transgenic mice in the absence of specific immunization presented with the same frequency of activated B cells as purposely immunized nontransgenic mice (n= 10, mean = 41.19%). compensate for the loss of CD19 function (6). We find that a viral protein A-9758 of a B-cell tropic human herpesvirus similarly modulates BCR signalingin vivo, resulting in hyperactive naive B cells. This viral protein is nuclear localized, and thus far most of its functionality has been attributed to transcriptional regulation and sequence-specific DNA binding. This protein is Rabbit Polyclonal to NFYC not an oncogene as defined by the classic, cell autonomous transformation assay. Ourin vivodata show that it is a subtler regulator of B-cell differentiation and suggest a novel mechanism as to how immunotropic viruses modulate BCR signaling to further the preferential survival and expansion of virally infected host cellsin vivo. Kaposi sarcoma-associated herpesvirus (KSHV) is a B-cell tropic human tumor virus. In addition to causing Kaposi sarcoma, it is also the causative agent of a type of post-germinal center (GC) diffuse large B-cell lymphoma (DLBCL), and primary effusion lymphoma (PEL), as well as the plasmablastic variant of multicentric Castleman’s disease (MCD) (9,10). KSHV can be found in peripheral blood, mature, CD19+B cells (15,37). During asymptomatic, latent persistence in CD19+B cells, as well as in 99% of PEL cells in culture, only the latency-associated nuclear antigen (LANA) and a few other viral latent genes in the LANA latency locus are expressed (18,20,27,45). We previously generated transgenic mice, which express LANA in CD19+mature B cells (21). In these mice LANA is under the control of its own viral promoter, which is B cell specific (28). LANA transgenic mice exhibit an activated mature B-cell phenotype, as evidenced by the expansion of CD19+IgM+IgD+FSChisplenic B cells. Functionally, B cells can be divided into follicular B cells (FL cells), which respond to T-cell-dependent antigens, and marginal zone B cells (MZ cells), which respond to multivalent, so-called T-cell-independent antigens. MZ cells also drive the primary (days 2 to 4) low-affinity B-cell response to antigen (reviewed in reference26). Both populations require BCR engagement and CD19 for proliferation and differentiation. FL cells require T-cell help in their response to monovalent antigens, e.g., the hapten 4-hydroxy-3-nitrophenylacetyl (NP). The B-cell-T-cell interaction takes place in the germinal centers (GC). Note that there are three steps in follicular B-cell activation: (i) antigen binding and transformation to activated B-cell blasts, which are FSChiand express the CD71 and CD68 activation markers (31); (ii) T-cell-dependent activation and migration into the GC; and (iii) T-cell-dependent A-9758 class switching, GC exit, and differentiation (reviewed in reference1). MZ B cells do not necessarily require T-cell help, although they do respond to T-cell-dependent antigens and are responsible for the primary IgM response (54). In addition, strong, multivalent antigens are thought to multimerize BCR molecules to such an extent that MZ cell activation is induced independently of direct T-cell contact (16,35). To test whether the LANA-induced hyperplasia is dependent on BCR-transmitted activation signals, we crossed LANA transgenic mice to isogenic CD19/mice (19,24,51). We found that the LANA transgene could not rescue the GC B-cell deficiency of CD19/mice. However, LANA was able to rescue the MZ defect in CD19/mice. We further found that LANA augmented the primary B-cell response to acute antigen, a finding consistent with a model in which LANA lowers the threshold for BCR-dependent B-cell A-9758 activation, proliferation, and survival. In the widest sense, thesein vivoobservations suggest that antigen exposure fosters latent persistence of B-cell-tropic herpesviruses, analogous to the way in which microbial translocations foster T-cell activation and HIV replication (8). Previously, it was reported that transgenic expression of the Epstein-Barr virus (EBV) LMP1 protein accelerated extrafollicular B-cell differentiationin vivomimicking CD40 signaling (60), that EBV LMP2A enhances B-cell responses (4,56), and that murine herpesvirus 68 (MHV68) coinfection fosters macrophage responses to bacterial antigens (7). The present study is the first demonstration that B cells expressing a KSHV viral protein displayed enhanced response to a T-dependent antigen. == MATERIALS AND METHODS == == Mice. == Transgenic mice expressing KSHV LANA were previously described (21). CD19 knockout mice [C.129P2-Cd19tm1(cre)Cgn/J] were purchased from the Jackson Laboratory (Bar Harbor, ME). LANA+/CD19 knockout mice were obtained by breeding LANA transgenic mice with CD19 knockout mice. Mice were maintained under pathogen-free conditions using ventilated cages. Animals were injected intraperitoneally with 100 g of 4-hydroxy-3-nitrophenylacetyl-keyhole limpet hemocyanin (NP-KLH; Biosearch Technologies, Inc., Novato, CA) in 200 l of phosphate-buffered saline (PBS). All experiments were approved by the Institutional Animal Care and Use Committee (IACUC) at the University of North Carolina at.