Abstract
BACKGROUND: The protein tyrosine phosphatase SHP-2 is known to regulate many cellular functions including growth, differentiation and survival. Interestingly, a gain-of-function mutation (residue E76K) in the gene encoding SHP-2 has been found in some colorectal cancers. Importantly, intestinal epithelial cell (IEC)-specific expression of SHP-2(E76K) mutant in mice was not sufficient to induce tumorigenesis but markedly promoted tumor growth under the Apc(Min/+) background. Conversely, SHP-2 silencing inhibited anchorage-independent growth of human CRC cells (Gagné-Sansfaçon et al., Oncotarget 2016). However, the molecular mechanisms involved in the pro-oncogenic action of SHP-2 remained to be identified. AIMS: To determine a possible role of the protein tyrosine phosphatase SHP-2 in the development of senescence. METHODS: Non immortalized human intestinal epithelial cells (HIEC) and human colorectal cancer cells (HCT116) were infected with lentiviruses encoding a shRNA directed against SHP-2. The impact on growth was assessed by BrdU incorporation, SA-β-galactosidase staining and qPCR or immunoblot analyses of regulatory proteins and signaling pathways regulating the cell cycle. SHP-2 deletion was induced ex vivo in intestinal organoids and the impact on organoid development was analyzed. RESULTS: SHP-2 silencing in HIEC changes cell morphology and results in the activation of the Wnt/β-catenin pathway and in the decreased activation of ERK1/2 MAP Kinases. Few days after silencing, the number of cells in S phase is significantly decreased and senescence (SA-β-galactosidase staining) as well as DNA damage (γH2AX) are observed. Western blot and qPCR analyses demonstrate increased expression of the cell cycle inhibitor p27 and increased phosphorylation of p53 on serine 15. Finally, deletion of SHP-2 in enteroids clearly limits their proliferative capacity and development; notably, few days after, organoids loose their integrity and degenerate in contrast to control organoids that continue to develop. CONCLUSIONS: In summary, our results suggest that SHP-2 protects intestinal epithelial cells against an oncogenic stress leading to senescence. FUNDING AGENCIES: CIHRCRS