Circular RNA CircHIPK3 Promotes Homeostasis of the Intestinal Epithelium by Reducing MicroRNA 29b Function

Gastroenterology. 2021 Oct;161(4):1303-1317.e3. doi: 10.1053/j.gastro.2021.05.060. Epub 2021 Jun 9.


Background & aims: Circular RNAs (circRNAs) are a class of endogenous noncoding RNAs that form covalently closed circles. Although circRNAs influence many biological processes, little is known about their role in intestinal epithelium homeostasis. We surveyed circRNAs required to maintain intestinal epithelial integrity and identified circular homeodomain-interacting protein kinase 3 (circHIPK3) as a major regulator of intestinal epithelial repair after acute injury.

Methods: Intestinal mucosal tissues were collected from mice exposed to cecal ligation and puncture for 48 hours and patients with inflammatory bowel diseases and sepsis. We isolated primary enterocytes from the small intestine of mice and derived intestinal organoids. The levels of circHIPK3 were silenced in intestinal epithelial cells (IECs) by transfection with small interfering RNAs targeting the circularization junction of circHIPK3 or elevated using a plasmid vector that overexpressed circHIPK3. Intestinal epithelial repair was examined in an in vitro injury model by removing part of the monolayer. The association of circHIPK3 with microRNA 29b (miR-29b) was determined by biotinylated RNA pull-down assays.

Results: Genome-wide profile analyses identified ∼300 circRNAs, including circHIPK3, differentially expressed in the intestinal mucosa of mice after cecal ligation and puncture relative to sham mice. Intestinal mucosa from patients with inflammatory bowel diseases and sepsis had reduced levels of circHIPK3. Increasing the levels of circHIPK3 enhanced intestinal epithelium repair after wounding, whereas circHIPK3 silencing repressed epithelial recovery. CircHIPK3 silencing also inhibited growth of IECs and intestinal organoids, and circHIPK3 overexpression promoted intestinal epithelium renewal in mice. Mechanistic studies revealed that circHIPK3 directly bound to miR-29b and inhibited miR-29 activity, thus increasing expression of Rac1, Cdc42, and cyclin B1 in IECs after wounding.

Conclusions: In studies of mice, IECs, and human tissues, our results indicate that circHIPK3 improves repair of the intestinal epithelium at least in part by reducing miR-29b availability.

Keywords: CircRNAs; IBD; IEC; MicroRNAs; Mucosal Injury.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Movement*
  • Cell Proliferation*
  • Cells, Cultured
  • Cyclin B1 / genetics
  • Cyclin B1 / metabolism
  • Disease Models, Animal
  • Down-Regulation
  • Epithelial Cells / metabolism*
  • Epithelial Cells / pathology
  • Female
  • Homeostasis
  • Humans
  • Inflammatory Bowel Diseases / genetics
  • Inflammatory Bowel Diseases / metabolism*
  • Inflammatory Bowel Diseases / pathology
  • Intestinal Mucosa / metabolism*
  • Intestinal Mucosa / pathology
  • Male
  • Mice, Inbred C57BL
  • MicroRNAs / genetics
  • MicroRNAs / metabolism*
  • Neuropeptides / genetics
  • Neuropeptides / metabolism
  • RNA, Circular / genetics
  • RNA, Circular / metabolism*
  • Sepsis / genetics
  • Sepsis / metabolism*
  • Sepsis / pathology
  • Wound Healing
  • cdc42 GTP-Binding Protein / genetics
  • cdc42 GTP-Binding Protein / metabolism
  • rac1 GTP-Binding Protein / genetics
  • rac1 GTP-Binding Protein / metabolism


  • Ccnb1 protein, mouse
  • Cdc42 protein, mouse
  • Cyclin B1
  • MIRN29 microRNA, mouse
  • MIRN29a microRNA, human
  • MicroRNAs
  • Neuropeptides
  • RNA, Circular
  • Rac1 protein, mouse
  • cdc42 GTP-Binding Protein
  • rac1 GTP-Binding Protein