Epithelial CRL4 Is Critical for Maintaining Intestinal Homeostasis Against DSS-Induced Colitis by Regulating the Proliferation and Repair of Intestinal Epithelial Cells.

BACKGROUND AND AIMS

Inflammatory bowel disease (IBD) is currently gaining an increasing global interest. Intestinal epithelial barrier dysfunction is crucial toward developing IBD; however, the underlying mechanisms are not yet elucidated. This study is aimed at elucidating the function of CRL4, an E3 ligase, toward mediating intestinal homeostasis.

METHODS

Colon samples were collected from patients with IBD and healthy individuals to examine the expression of CRL4. CRL4 conditional knockdown in mouse intestinal epithelial cells (IECs) (DCAF2) were constructed. DCAF2 and their littermate control (DCAF2) were treated with dextran sodium sulfate (DSS) to induce acute colitis. Transcriptome analysis was performed on inflamed colon samples obtained from the mice. Cell cycle regulators were evaluated using real-time polymerase chain reaction (PCR), while tight junction and apoptosis proteins were examined via immunofluorescence and western blot.

RESULTS

CRL4 expression was significantly decreased in the inflamed IBD epithelium, and low expression of CRL4 associated with high recurrence risk. Mice with DCAF2 specific knockout in IECs suffer from embryonic death. Multiple genes involved in cell proliferation, immune response, and gap junction were differentially expressed in inflamed colon from DCAF2 compared with DCAF2. Furthermore, conditional downregulation of CRL4 in the intestinal epithelium induced primarily epithelial damage, increased intestinal permeability, and diminished tight junction protein expression. In vivo and in vitro cell transfection experiments revealed that CRL4 enhanced cell proliferation by promoting p21 ubiquitination and degradation, thereby inhibiting G2/M cell cycle. In addition, CRL4 can also inhibit IEC apoptosis and promote cell autophagy.

CONCLUSIONS

CRL4 downregulation in IECs promotes intestinal barrier dysfunction and inhibits IEC proliferation, thus making it more susceptible to inflammation.