Stable PTPN2 knockdown in human intestinal epithelial cell lines increases claudin‐2 expression and activation of MEK/ERK and PI3K signaling (1111.7)

Abstract

Stable PTPN2 Knockdown in Human Intestinal Epithelial Cell Lines increases Claudin‐2 expression and activation of MEK/ERK and PI3K signaling Moorthy Krishnan and Declan F McCole, Division of Biomedical Sciences, University of California, Riverside, Riverside, CA 92521.Genome‐wide association studies identified protein tyrosine phosphatase non‐receptor type‐2 (PTPN2) as a susceptibility gene for inflammatory bowel disease (IBD), Diabetes and Celiac disease. Increased permeability of the intestinal epithelium is believed to be an important factor contributing to IBD. The aim of this study was to generate a stable PTPN2‐deficient intestinal epithelial cell line to investigate the role of PTPN2 and its protein product, T‐cell protein tyrosine phosphatase (TCPTP), in protecting epithelial barrier function. We developed Caco2‐BBE and HT‐29 intestinal epithelial cell lines (IECs) stably expressing short hairpin RNA for PTPN2. Western blotting and densitometry indicated that TCPTP protein expression was reduced by 50 ± 0.3% in PTPN2‐depleted Caco2‐BBE and HT‐29 cells, while there was a 3.5 ± 1.3 and 4.2 ± 1.5 fold increased expression of claudin‐2 in Caco2‐BBE and HT‐29 cells respectively. Furthermore, knockdown of PTPN2 resulted in decreased transepithelial electrical resistance (TER) (76 ± 3% and 50 ± 4%) in Caco2‐BBE and HT‐29 cells respectively) compared to control, and this correlated with the observed increase in claudin‐2 expression. Interestingly, Western blot and immunofluorescence studies showed that basal phosphorylation of ERK1/2 and Akt was increased in PTPN2 KD cells. In conclusion, we report an inverse relationship between PTPN2 (TCPTP) and claudin‐2 expression associated with increased activation of MEK/ERK and PI3K pathways in stable PTPN2‐deficient human IECs.Grant Funding Source: Supported by NIH R01DK091281