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Updated: Jun 26, 2025
Monitoring Hippo Signaling Pathway Activity Using a Luciferase-based Large Tumor Suppressor LATS Biosensor
Published on: September 13, 2018
Bcl-2 dependent modulation of Hippo pathway in cancer cells
Simona D'Aguanno1, Matteo Brignone2, Stefano Scalera3
1Preclinical Models and New Therapeutic Agents Unit, IRCCS Regina Elena National Cancer Institute, Rome, 00144, Italy. simona.daguanno@ifo.it.
Introduction:
Bcl-2 and Bcl-xL are the most studied anti-apoptotic members of Bcl-2 family proteins. We previously characterized both of them, not only for their role in regulating apoptosis and resistance to therapy in cancer cells, but also for their non-canonical functions, mainly including promotion of cancer progression, metastatization, angiogenesis, and involvement in the crosstalk among cancer cells and components of the tumor microenvironment. Our goal was to identify transcriptional signature and novel cellular pathways specifically modulated by Bcl-2.
Methods:
We performed RNAseq analysis of siRNA-mediated transient knockdown of Bcl-2 or Bcl-xL in human melanoma cells and gene ontology analysis to identify a specific Bcl-2 transcriptional signature. Expression of genes modulated by Bcl-2 and associated to Hippo pathway were validated in human melanoma, breast adenocarcinoma and non-small cell lung cancer cell lines by qRT-PCR. Western blotting analysis were performed to analyse protein expression of upstream regulators of YAP and in relation to different level of Bcl-2 protein. The effects of YAP silencing in Bcl-2 overexpressing cancer cells were evaluated in migration and cell viability assays in relation to different stiffness conditions. In vitro wound healing assays and co-cultures were used to evaluate cancer-specific Bcl-2 ability to activate fibroblasts.
Results:
We demonstrated the Bcl-2-dependent modulation of Hippo Pathway in cancer cell lines from different tumor types by acting on upstream YAP regulators. YAP inhibition abolished the ability of Bcl-2 to increase tumor cell migration and proliferation on high stiffness condition of culture, to stimulate in vitro fibroblasts migration and to induce fibroblasts activation.
Conclusions:
We discovered that Bcl-2 regulates the Hippo pathway in different tumor types, promoting cell migration, adaptation to higher stiffness culture condition and fibroblast activation. Our data indicate that Bcl-2 inhibitors should be further investigated to counteract cancer-promoting mechanisms.
Insights
Bcl-2 protein regulates the Hippo pathway, impacting cancer cell migration and fibroblast activation. Targeting Bcl-2 may offer new cancer treatment strategies by counteracting these cancer-promoting mechanisms.
Area of Science:
- Molecular Biology
- Cancer Biology
- Cellular Signaling
Background:
- Bcl-2 and Bcl-xL are key anti-apoptotic proteins with known roles in cancer therapy resistance.
- Beyond apoptosis, these proteins also promote cancer progression, metastasis, and angiogenesis.
- Their involvement in tumor microenvironment crosstalk is significant, necessitating further investigation into their specific functions.
Purpose of the Study:
- To identify the unique transcriptional signature modulated by Bcl-2.
- To uncover novel cellular pathways specifically regulated by Bcl-2.
- To elucidate the non-canonical functions of Bcl-2 in cancer progression.
Main Methods:
- RNA sequencing and gene ontology analysis were used to identify Bcl-2's transcriptional signature.
- Validation of Hippo pathway gene expression via qRT-PCR in multiple cancer cell lines.
- Western blotting, YAP silencing, migration assays, and co-culture experiments assessed Bcl-2's functional impact.
Main Results:
- Bcl-2 was shown to modulate the Hippo Pathway by affecting upstream YAP regulators across various cancer types.
- YAP inhibition abrogated Bcl-2's pro-migratory and pro-proliferative effects on cancer cells, especially under stiff conditions.
- Bcl-2's ability to stimulate fibroblast migration and activation was dependent on YAP.
Conclusions:
- Bcl-2 plays a crucial role in regulating the Hippo pathway, influencing cancer cell migration and adaptation to mechanical stress.
- Bcl-2 promotes cancer progression through activation of fibroblasts within the tumor microenvironment.
- Bcl-2 inhibitors represent a promising therapeutic avenue to disrupt these cancer-promoting functions.
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