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Updated: Oct 19, 2025
Modeling Paracrine Noncanonical Wnt Signaling In Vitro
Published on: December 10, 2021
A CRISPR knockout screen reveals new regulators of canonical Wnt signaling
Tamar Evron1, Michal Caspi1, Michal Kazelnik1
1Department of Clinical Microbiology and Immunology, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel.
Abstract:
The Wnt signaling pathways play fundamental roles during both development and adult homeostasis. Aberrant activation of the canonical Wnt signal transduction pathway is involved in many diseases including cancer, and is especially implicated in the development and progression of colorectal cancer. Although extensively studied, new genes, mechanisms and regulatory modulators involved in Wnt signaling activation or silencing are still being discovered. Here we applied a genome-scale CRISPR-Cas9 knockout (KO) screen based on Wnt signaling induced cell survival to reveal new inhibitors of the oncogenic, canonical Wnt pathway. We have identified several potential Wnt signaling inhibitors and have characterized the effects of the initiation factor DExH-box protein 29 (DHX29) on the Wnt cascade. We show that KO of DHX29 activates the Wnt pathway leading to upregulation of the Wnt target gene cyclin-D1, while overexpression of DHX29 inhibits the pathway. Together, our data indicate that DHX29 may function as a new canonical Wnt signaling tumor suppressor and demonstrates that this screening approach can be used as a strategy for rapid identification of novel Wnt signaling modulators.
Insights
Researchers discovered DHX29 as a novel Wnt signaling inhibitor. Knocking out DHX29 activates the Wnt pathway, while its overexpression inhibits it, suggesting DHX29 acts as a tumor suppressor in colorectal cancer.
Area of Science:
- Molecular Biology
- Genetics
- Oncology
Background:
- Wnt signaling pathways are crucial for development and homeostasis.
- Aberrant Wnt signaling, particularly the canonical pathway, drives diseases like colorectal cancer.
- Identifying novel regulators of Wnt signaling is essential for therapeutic development.
Purpose of the Study:
- To identify new inhibitors of the oncogenic canonical Wnt pathway using a genome-wide CRISPR-Cas9 screen.
- To characterize the role of DExH-box protein 29 (DHX29) in regulating Wnt signaling.
Main Methods:
- Genome-scale CRISPR-Cas9 knockout screen.
- Assay based on Wnt signaling-induced cell survival.
- Analysis of Wnt target gene expression (e.g., cyclin-D1).
- Investigating the effects of DHX29 knockout and overexpression.
Main Results:
- A genome-wide CRISPR-Cas9 screen identified novel Wnt pathway inhibitors.
- Knockout of DHX29 activates the canonical Wnt pathway.
- DHX29 knockout leads to upregulation of the Wnt target gene cyclin-D1.
- Overexpression of DHX29 inhibits the Wnt pathway.
Conclusions:
- DHX29 functions as a novel tumor suppressor in the context of canonical Wnt signaling.
- The CRISPR-Cas9 screening approach is effective for rapid identification of Wnt signaling modulators.
- DHX29 is a potential therapeutic target for Wnt-driven cancers, including colorectal cancer.
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