? IL-27-producing B cells in the antibody response Built on our novel findings that B cells can be induced to express high levels of IL-27 (IL27p28/EBI3 heterodimer), this proposal will explore molecular and cellular mechanisms underlying the role of IL-27 and IL-27-producing B cells (B-27 cells) in class-switched antibody (Ab) responses. As we contend, B-27 cells shape the magnitude and quality of T-dependent Ab responses, including those elicited by viral infections. As we also contend, they do so as ?helper? B cells to enhance the function of their target ?effector? B cells, i.e., those responding to antigens and differentiating into IgG-producing cells. Such class-switched Abs include IgG1, which directly neutralizes virus, and human IgG3 and mouse IgG2a/IgG2c (IgG2a), which have additional anti-viral effector functions. Owing to the unique requirement of both innate and adaptive immune receptor signals for their induction, B-27 cells are strategically positioned to mediate the potent effect of TLR ligands in boosting the Ab response. We hypothesize that B-27 cells are induced in a manner dependent on transcription factor BATF3, and optimize the Ab response by cooperating with IFNg to promote proliferation, survival and full differentiation of IgG-producing B cells. This is based on our compelling preliminary data indicating that: (i) as an important source of IL-27, B cells are induced to produce IL-27 after priming by TLR ligands and then stimulation by Tfh cell stimuli CD154 and IL-21; (ii) mice with B cell-specific deficiency in Il27p28, Ebi3 or Batf3 are impaired in IL-27 production and specific IgG2a responses; (iii) IL-27 activates STATs, directs CSR to IgG2a and promotes survival and plasma cell differentiation in B cells stimulated by CD154 and IL-21 in vitro; and (iv) IL-27 and IFNg together boost B cell growth and differentiation in vitro, and, conversely, combined IL-27R and IFNgR deficiency in B cells abrogates specific IgG2a responses and significantly impairs IgG1 responses in vivo. To test our hypothesis, we will (Aim 1) characterize B-27 cells induced in Tg(Il27p28-Gfp) reporter mice upon infection by vaccinia virus (VV), which is also used as a vector of a variety of vaccines, or immunization with conjugated hapten NP-CGG mixed with alum and TLR ligand LPS; and determine the cooperation of B-27 cells with IFNg in specific IgG responses and underlying effector B cell proliferation, survival, CSR/SHM, plasma cell and memory B cell differentiation. We will also (Aim 2) address the mechanisms underlying the role of BATF3 in Il27p28 induction by identifying its cis-elements in the Il27p28 locus and partner transcription factors in vitro; and analyze induction of B cell BATF3 and its impact on B-27 cell generation and Ab responses to VV infection and NP-CGG/alum/LPS in vivo. Finally, we will (Aim 3) analyze the cooperation of B-cell IL-27R and IFNgR signals in optimizing effector B cell growth and differentiation in response to VV infection in vivo and stimulation with CD154 and IL-21 in vitro; and address the underlying mechanisms, focusing on their synergy in regulating STAT signal outputs and gene expression. By unveiling the mechanisms and function of B-27 cells, our studies will have a sustained impact on the understanding of IgG responses to viral infections and vaccine development.
As one of the most successful medical interventions in human history, vaccines elicit antibodies to afford immune protection that has eradicated small pox and all but eradicated polio, diphtheria, tetanus, mumps, and measles from the developed world, transforming public health practices in the process. To explore mechanisms underlying anti-viral antibody responses towards development of sophisticated vaccine strategies to deal with recalcitrant infectious agents, here we will analyze the induction of a highly specialized subset of B lymphocytes, i.e., those producing cytokine IL-27 with pleiotropic functions, by vaccinia virus, which was the small pox vaccine and is currently used as vectors of several vaccines (including the HIV vaccine in clinical trial and a recent SARS-CoV-2 vaccine). We will also address the role of IL-27-producing B cells in optimizing protective anti-viral antibody response and underlying mechanisms, including how such B cells to promote the growth and differentiation of antibody-producing B cells, thereby opening a new avenue of research in the vaccine development.
