Fig. 6

An outer membrane protein of A. muciniphila Amuc_1100 coupled with host CREBH to regulate IEC proliferation and gut barrier integrity. IPEC-J2 or Caco2 cells were transfected with empty vector (pcDNA3.1) or vector expressing Amuc_1100 (Amuc) for 48 h. Total mRNA or protein was extracted for the following analyses: A Immunoblot analysis of Amuc_1100 (Amuc), Full-length CREBH, N-terminal CREBH (N-CREBH, active form), and β-actin. B mRNA expression of TLR2 and TLR4. C Immunoblot analysis of Grp78, eIF2α-p, CLDN2, and β-actin. D mRNA expression of CREBH, IGF1R, IGFBP5, CLDN5, and CLDN8 in the transfected IPEC-J2 cells. E Immunofluorescent confocal images of the transfected IPEC-J2 cells immune-stained with anti-CREBH and anti-TARF6 antibodies, and Hoescht 33,349 as described in the Additional file 1: Supplemental Methods. F Overlap coefficient analysis of the immunofluorescence images in E, showing co-localisation of CREBH and TRAF6. G Immunoblot analysis of Amuc_1100 (Amuc) and β-actin expression in Caco2 cells transfected with empty vector or Amuc_1100 (Amuc) expressing vector (for 48 h) treated with or without TNFα (20 ng/mL) for 24 h. Results represent the means with SD. The two-tailed Student’s t-test was used for statistical analyses of two-group comparisons. *P < 0.05 and **P < 0.01 versus controls