804 Genetic dissection of TLR9 reveals complex regulatory and cryptic pro-inflammatory roles in murine lupus

In lupus, TLR7 and TLR9 mediate loss of tolerance to RNA and DNA, respectively. Yet, TLR7 promotes while TLR9 protects from disease, implying differences in signaling. To dissect this 'TLR paradox', we generated two TLR9 point mutants—lacking either ligand (TLR9K51E) or MyD88 (TLR9P915H) binding—in lupus-prone MRL/lpr mice. Ameliorated disease of Tlr9K51E mice compared to Tlr9-/- controls revealed a TLR9 'scaffold' protective function that is ligand- and MyD88-independent. Unexpectedly, Tlr9P915H mice were more protected than both Tlr9K51E and Tlr9WT mice, suggesting that TLR9 also possesses ligand-dependent, but MyD88-independent, regulatory signaling and MyD88-mediated proinflammatory signaling. Triple mixed bone marrow chimeras showed that TLR9-MyD88 independent regulatory roles were B cell-intrinsic and restrained differentiation into pathogenic age-associated B cells (ABC) and plasmablasts (PB). These studies reveal MyD88-independent regulatory roles of TLR9, shedding light on the biology of endosomal TLRs. In addition, we have created TLR7/TLR9 chimeric molecules in the germline of MRL/lpr lupus-prone mice, in which the TIR domains have been switched, and determined the effect on disease outcomes. In this way, we could map the differential regulatory and proinflammatory functions of these two distinct endosomal TLRs. This work was supported by NIH grant R37-AI118841.