Given the possibility of crosstalk between TLRs, if expression of TLRs in the ileum was altered in response to norovirus infection, TLR4 response to γ-PGA would also be affected. The ileum with PPs of norovirus-infected mice showed no significant increase in TLR4 expression. Expression levels of TLR2, TLR3, TLR6, TLR8, and TLR9 were slightly increased. Though the increase was less than 2-fold, it was statistically significant (Fig. 6a). The expression of TLR1, which recognizes peptidoglycan and lipoproteins43,44, and TLR5, which recognizes bacterial flagellin45 was upregulated by ~5-fold and 2.5-fold, respectively, suggesting the possibility of crosstalk between TLR signaling through some of these upregulated TLRs. This might have contributed to the amplification of γ-PGA-mediated TLR4 signaling in vivo as several TLR downstream signaling pathways share transcription factors such as NF-κB and IRF-335, used for type I IFN production. In MLNs, expression levels of TLR2, TLR4, TLR6, TLR7, and TLR8 were slightly decreased (less than 2-fold) (Fig. 6b). These results suggest differentially altered TLR expression profile for norovirus infection in the ileum with PPs and MLNs in vivo. Interestingly, this expression profile of TLRs was different from that seen in virus-infected RAW264.7 cells (Fig. 2F), suggesting the involvement of commensal microbiota in the regulation of TLR expression in the ileum, in the context of norovirus infection in mice.Figure 6Impact of norovirus infection on TLR expression in mice. The ileum with PPs (a) and the MLNs (b) from BALB/c mice (6 weeks old, n = 9–10 per group) infected with MNV-1 as in Fig. 5 were retrieved 2 days after viral infection and expression of TLR mRNA was quantified as described in Fig. 2f. Data are represented as mean ± SD. *P < 0.05; **P < 0.01; ***P < 0.001; n.s., not significant; by two-tailed unpaired Student’s t-test.