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. Author manuscript; available in PMC: 2018 Mar 1.
Published in final edited form as: Gastroenterology. 2016 Nov 19;152(4):851–866.e24. doi: 10.1053/j.gastro.2016.11.018

Figure 6. F nucleatum regulates miR21 expression through TLR4/MYD88/NFκB pathway.

Figure 6

A and B) Knockdown of p65 in HCT116 or LoVo significantly down-regulated miR21 expression (**P < 0.01 by unpaired Student’s t test. Bars represent SD of three experiments). C) We generated luciferase reporter plasmids containing either wild type (WT) or mutant type (MT) of p65 binding sites in miR21 promoter. HEK293T cells were transfected with p65 shRNA or luciferase constructs (pGL3-WT and pGL3-MT) (**P < 0.01 by unpaired Student’s t test). D and E) HCT116 cells and LoVo were treated with F nucleatum or PBS following ChIP assay. qPCR results showed a more than 5 fold enrichment of miR21 promoter in p65 pulled-down DNA samples compared to IgG samples (**P < 0.01 by unpaired Student’s t test). F and G) The qRT-PCR assay indicated that F nucleatum failed to up-regulate miR21 expression when TLR4 or MYD88 was silenced in HCT116 and LoVo (**P < 0.01 by unpaired Student’s t test). H) Western blot analysis showed that CRC tissues from F nucleatum-treated APCmin/+ mice showed activation of NFκB, as well as other members including RASA1, PDCD4 and MAPK pathway (n = 4 mice per group).