Deciphering TLR3 and TLR7-mediated control of extrafollicular B cell proliferation and plasma cell differentiation 4202

Description Extrafollicular (EF) B cell responses are essential for protective immunity, but the signals that direct naïve B cells toward an EF fate remain unclear. Our previous work showed that deleting both adaptor proteins, MyD88 and TRIF, reduced EF cell numbers post-influenza infection. In separate studies, we show that TLR3 activation appears to be upstream of TRIF and TLR7 activation upstream of MyD88 signaling, but what role each of these adaptor proteins play in EF formation remains undefined. To address this, we performed flow cytometric analysis of CTV-labeled naïve B cells stimulated with TLR3, TLR7, or combined TLR3 + 7 agonists over a 96-hour period. Stimulation through TLR7 (CL097) promoted robust proliferation without plasma-cell differentiation, while TLR3 stimulation (POLY(I:C)) induced activation, as measured by increased cell surface expression of co-stimulatory receptors. Notably, TLR3 + 7 co-stimulation synergistically enhanced both proliferation and plasma-cell differentiation. qPCR analysis of key plasma-cell differentiation factors, including Pax5, Bach2, BLIMP1 and IRF4, all of which were strongly upregulated by combined TLR3 + 7 stimulation. To further dissect the transcriptional targets downstream of each TLR signal alone or in combination, we are using bulk RNA-Seq analysis with the goal to delineate the signaling pathways that explain the synergistic effects of these two distinct TLR stimuli on the kinetics of plasma cell differentiation in vitro. Funding Sources Funding sources: R01AI148652 and R01AI184867 Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)