Bachmann and Zinkernagel, 1997; Fehr et al., 1998). In con­ trast to TI antigens, TD antigens are generally monomeric soluble proteins that display single or few epitopes to antigen­ specific B cells and require cognate T cell help for induction of highly specific antibody responses generated through ger­ minal center reactions. Although not absolute, a general division of labor is also acknowledged between B cell subsets and the response to TI­ 2 and TD antigens. B1 and MZ B cell subpopulations have been considered to be primarily responsible for the antibody response to TI­2 antigens (Fagarasan and Honjo, 2000; Martin et al., 2001; Balázs et al., 2002), whereas FO B cells dominate antibody responses to soluble protein TD antigens. In accord with generating rapid antibody responses, MZ and B1 B cells have lower thresholds for activation com­ pared with FO B cells and are physically poised at sites either in tissues or at the blood–lymphoid interface that facilitates these early responses (Martin and Kearney, 2002). B1 and MZ B cells are described as innate B cells in that they express a restricted repertoire of germline­encoded BCRs with poly ­ reactive specificities (Bendelac et al., 2001). Responding MZ B cells produce antigen­ specific antibody at extrafollicular splenic sites early during the antibody response that is low af­ finity and predominantly IgM but also includes limited IgG subclasses. Although evidence exists that MZ B cells can also mount TD responses and initiate germinal center reactions (Song and Cerny, 2003; Phan et al., 2005), the ability of FO B cells to directly participate in rapid extrafollicular TI­2 anti­ body responses is minor (Ron and Sprent, 1985; Goud et al., 1988; Liu et al., 1988). Characterization of the TI­ 2 antibody response has predominantly relied on hapten­ coupled carbohydrates as model antigens. However, physiological TI­ 2 antigens would rarely be encountered in isolation but, rather, are typically associated with pathogen­ associated molecular patterns (PAMPs) recognized by pattern recognition re­ ceptors (PRRs; Janeway, 1989; Snapper, 2006). TLRs are one family of innate immune PRRs that have been shown to enhance the antibody response (Coutinho et al., 1974; Seppälä and Mäkelä, 1984; Sen et al., 2005; Heer et al., 2007; Rubtsov et al., 2008; Eckl­ Dorna and Batista, 2009). However, whether this regulation is physiologically rele­ vant has been controversial (Pasare and Medzhitov, 2005; Gavin et al., 2006). A major consequence of signaling by many PRRs is the rapid elaboration of inflammatory type I IFN cytokines (Baccala et al., 2007; McCartney and Colonna, 2009), and many viral and bacterial infections lead to IFN­ / produc­ tion (Bogdan, 2000; Mancuso et al., 2009). Type I IFN has been previously characterized to modulate antibody production both in vitro and in vivo, including promoting class­switch recombination and polarizing antibody responses toward IgG2a/c production (Finkelman et al., 1991; Le Bon et al., 2001, 2006; Coro et al., 2006; Heer et al., 2007). How IFN­ / acts to effect these changes in humoral immunity is not well established, nor is whether the type I IFN produced as a 1486 result of innate immune receptor signaling contributes to regulating the antibody response. It has been proposed that in addition to their antigen­ specific BCR, B cells may also use innate mechanisms for pathogen recognition. However, it is still unclear which of the innate receptors are used by B cells or what the conse­ quence of these signals is to the B cell antibody response. The rapid TI­2 antibody response provides a temporal bridge be ­ tween the early innate and late adaptive immune responses. Thus, we were interested to investigate how innate signals would augment the TI­2 antibody response and influence the different B cell populations during this response. In this paper, we show that TLR agonists significantly enhance the TI­2 antibody response through the elaboration of type I IFN that promotes production of IgG2c by FO B cells. This contribution elicited by FO B cells is shown to accelerate the kinetics and enhance the magnitude and qual­ ity of the TI­2 antibody response. Thus, these data not only provide a mechanistic example of how innate immunity influences adaptive immunity but also define how humoral immunity counters physiological pathogens that present TI­ 2 antigens in the presence of ligands for innate immune receptors. RESULTS Poly(I:C) enhances the magnitude and kinetics of the TI-2 antibody response Poly(I:C) is a synthetic double­stranded RNA (dsRNA) that mimics viral nucleic acids and is recognized by several innate PRRs, including the TLR3 receptor. MZ B cells are consid­ ered to be largely responsible for the antibody response to TI­2 antigens (Martin et al., 2001) and express significantly higher levels of the TLR3 transcript than FO B cells (Genestier et al., 2007). Thus, we questioned how poly(I:C) might in­ fluence the TI­2 antibody response by immunizing mice with the model TI­2 antigen NP­Ficoll in the presence or absence of poly(I:C). These results demonstrated that poly(I:C) markedly enhanced the antigen­ specific IgG3 and IgG2c antibody responses at both day 4 and day 7 after immuniza­ tion (Fig. 1 A). In contrast, poly(I:C) did not alter the day­ 4 IgM response but instead promoted a significant decrease in the NP­specific IgM response of 1.6­fold at 7 d after immu ­ nization (Fig. 1 A). Overall, poly(I:C) led to a twofold increase in total antigen­specific Ig 7 d after immunization (Fig. 1 B). The composition of the TI­2 antigen­specific antibody response was skewed by poly(I:C) from an IgM­dominated (61.3%) to an IgG­dominated (81.6%) response (Fig. 1 B). However, poly(I:C) did not similarly enhance the magnitudes of all IgG subclasses but, rather, had the most striking effect on the NP­specific IgG2c response by accelerating the kinetics and increasing the magnitude of this isotype (Fig. 1, A and B). Relative to other IgG isotypes, IgG2c is the least abundant isotype produced during the NP­ Ficoll response in the absence of poly(I:C) (Fig. 1 B; Mongini et al., 1981, 1984). Enhancement of the IgG2c response by poly(I:C) was dose dependent (Fig. S1 A) and, importantly, not a result of IFN- drives FO B cell TI-2 responses | Swanson et al.