Mutations In The DNA-binding Domain Of FXR Impaired Intestinal Anti-inflammation And Lipid Homeostasis

The farnesoid X receptor(FXR)is a transcription factor which acts through ligand binding and belongs to nuclear receptor superfamily.Highly expressed in liver.intestine,kidney and adrenals,FXR plays a pivotal role in bile acid homeostasis,lipid metabolism and inflammation regulating.FXR also became a hot target in drug discovery on account of its satisfying therapeutic effect in non-alcoholic fatty liver,type 2 diabetes,ulcerative colitis,atherosclerosis,cholestasis and other diseases.However,the research and drug discovery targeting FXR mainly focused on its transcriptional regulation,through activating or inhibiting the transcription of target genes.Non-transcriptional regulatory mechanism remains poorly studied due to the lack of suitable research tools.Starting from structural biology,we found three decisive sites,which are Arg 152,Arg 153,and Lys 157,can affect the interaction between FXR DNA binding domain and its response element,IR-1(inverted repeats-1).We further verified this through AlphaScreen assay in vitro and cell based reporter assay in vivo,which all showed that mutations to alanine in the three sites destroyed the interaction between FXR DBD and DNA.Next,we constructed genetically modified mice with three mutations in FXR DBD,and further verified the loss of transcriptional activity through detecting the expression of FXR target genes.Through comparing the phenotype of wild type mice and mutation mice in normal feeding conditions,high fat diet(HFD)-induced obesity conditions,and dextran sulfate sodium(DSS)-induced ulcerative colitis conditions,we found that the mutations impaired mice intestinal anti-inflammation and lipid homeostasis.What’s more,we found the non-transcriptional regulation in FXR plays a leading role in maintaining bile acid homeostasis,and a partial role in regulating lipid metabolism.In contrast,the intestinal anti-inflammatory regulation by FXR is only regulated through its transcriptional pathway.This is the first time that a mouse model has been used to study transcriptional and non-transcriptional regulation mechanism of FXR,which also provides new ideas in the development and evaluation of drugs targeting FXR.