The stem cell ubiquitin ligase RNF43 in gastric homeostasis, DNA damage response and carcinogenesis: from biological function to application as biomarker
Cell Biology
Final Report Abstract
The E3 ubiquitin ligase RING finger protein 43 (RNF43) has been reported to be an important player in gastrointestinal carcinogenesis and it was shown to be mutated in many different cancer types. Mutations in RNF43 were described as driving mutations in gastric cancer key in the transition from adenomas to carcinomas. The tumor suppressor activity of RNF43 in colon and pancreas was related to its Wnt-inhibitory capacity. However, the role of RNF43 in the stomach was unclear and therefore was one of the main focuses of this study. Using cells engineered by CRISPR/CAS9 or transduced by shRNF43 lentiviruses, we observed that loss of RNF43 function enhanced the tumorigenic potential of these cells not only in vitro but also in vivo using a xenograft mouse model. In these tumors, we observed that depletion of RNF43 correlated with higher levels of the gastric stem cell markers, while no changes in Wnt activation was detected. The tumor suppressor function of endogenous RNF43 was further supported by two mouse models generated in our lab (Rnf43ΔEx8 and Rnf43H292R/H295R). Multifocal hyperplasia and thickening of the gastric mucosa, which increased over time, was observed in these mice. However, these changes were not related to alterations in the Wnt pathway and prompted us to explore other signaling pathways that could explain the tumor suppressive function of RNF43 in the stomach. Considering the involvement of other RING finger proteins in the DDR, we explored whether RNF43 was also involved in DDR in the stomach. Cells lacking functional RNF43 showed reduced activation of DDR and apoptosis upon irradiation. Moreover, these cells were more resistant to chemotherapeutics inducing DNA not only in vivo but also in vivo. In addition, RNF43 expression was correlated to susceptibility to chemotherapy of human gastric organoids. Since H. pylori infection is the major risk factor for gastric cancer development, and previous studies showed that H. pylori directly induces DNA damage in gastric epithelial cells, we investigated how loss of RNF43 influenced the course of infection. Rnf43ΔEx8 mice infected for 3 months showed higher inflammation and lower activation of DDR and apoptosis compared to wild type mice. Higher inflammation and development of metaplasia was detected in long-term infected Rnf43H292R/H295R mice, confirming that loss of Rnf43 function aggravates H. pylori-driven pathology. Taken together, our results confirm a tumor suppressor function of RNF43 in the stomach independent of Wnt signaling pathway and suggest that the analysis of RNF43 expression and mutation analysis may be used as biomarkers for therapy response.
Publications
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Loss of endogenous RNF43 function enhances proliferation and tumour growth of intestinal and gastric cells. Carcinogenesis. 2019 Jun 10;40(4):551-559
Neumeyer V, Grandl M, Dietl A, Brutau-Abia A, Allgäuer M, Kalali B, Zhang Y, Pan KF, Steiger K, Vieth M, Anton M, Mejías-Luque R, Gerhard M
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Mutated Rnf43 Aggravates Helicobacter Pylori- Induced Gastric Pathology. Cancers (Basel). 2019 Mar 16;11(3):372
Neumeyer V, Vieth M, Gerhard M, Mejías-Luque R
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Loss of RNF43 function contributes to gastric carcinogenesis by impairing DNA damage response. Cell Mol Gastroenterol Hepatol. 2020 Nov 11:S2352-345X(20)30184-3
Neumeyer V, Brutau-Abia A, Allgäuer M, Pfarr N, Weichert W, Falkeis-Veits C, Kremmer E, Vieth M, Gerhard M, Mejías-Luque R
