UHRF1 Mediated DNA Methylation Is Required For Intestinal Homeostasis And Regeneration

DNA methylation is an important epigenetic modification,which is catalyzed by DNA methyltransferase and can affect cell fate through regulation of gene transcription.As development progresses and lineage choice decisions are made,many loci undergo cell-type-specific DNA methylation changes.Our understanding of the role of DNA methylation in tissue development and homeostasis is still not complete,as well as its role in the tissue damage repair process.How and to what degree DNA methylation affects stem cell self-renewal and differentiation are poorly understood.It has been reported that UHRF1,as a multidomain protein,can recognize and directly bind to the hemimethylated DNA and methylated histone H3,then recruit DNMT1 to the replication fork to copy methylation pattern during DNA replication.As known biochemical function of UHRF1,the physiological role of UHRF1-mediated DNA methylation in vivo has attracted more attention but its physiological role in animals are largely unknown.The tight cell hierarchy and rapid renewal rate of the intestinal epithelium make it an ideal model system for studying stem cells in vivo.Through establishing intestinal specific conditional knockout and histone binding site mutation(YP187/188AA)Uhrf1 mouse model,we carried out the research on the function and mechanism of Uhrf1 mediated DNA methylation in intestinal development and stem cell homeostasis.We found that Uhrf1 is mainly expressed in proliferative cells(stem and progenitor cells)of intestinal epithelium.Embryonic knockout of Uhrf1 blocked early intestinal development,and broke the balance of intestinal homeostasis,accompanied weight loss and death of the mice.However,the intestinal epithelium is apparently normal in histone binding site mutation mouse.Then we knocking-out Uhrf1 in adult mice,there is no apparent morphological change of epithelial in the short term,butthe homostasis of intestinal severely disturbed in the ong run,with decreased cell proliferation and differentiation and increased cell apoptosis.In addition,Uhrf1 is essential for the regeneration process after X-Ray radiation-induced epithelial damage.Mechanically,data shown that ablation of Uhrf1 lead to DNA hypomethylation,DNA damage arising and cell cycle arrest.Notably,cell cycle suppressor Cdkn1a(p21)gene expression was significantly up-regulated in the crypt region.Bisulfite sequencing analysis was then performed to assess the DNA methylation status of Cdkn1 a,and we found that Cp G sites of Cdkn1 a gene showed a decreased DNA methylation level in Uhrf1-deficient intestinal cells than in WT.This decreased DNA methylation potentially led to increased p21 expression,causing cell cycle arrest and apoptosis.Based on above work,we suggest that DNA methylation mediated by Uhrf1 can maintain the homeostasis of intestinal epithelium,promote regeneration and regulate stem cell self-renewal and differentiation through inhibiting the expression of cell cycle suppressor p21.Our study will extend current understanding of the physiological function of DNA methylation mediated by UHRF1 and uncover epigenetic regulation mechanism of stem cell homeostasis controlled by DNA methylation.