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Non-canonical Notch Signaling Regulates Actin Remodeling in Cell Migration by Activating PI3K/AKT/Cdc42 Pathway
Lei Liu1,2,3, Lin Zhang1,2, Shuo Zhao4
1Department of Biochemistry and Molecular Biology, Nanjing Medical University, Nanjing, China.
Abstract:
Tumor cell migration is a critical step in cancer metastasis. Over-activated Notch pathway can promote the migration of cancer cells, especially in the breast cancer. However, the underlying mechanism of non-canonical Notch signaling in modulating the migration has not yet been clearly characterized. Here we demonstrated that DAPT, a gamma secretase inhibitor, inhibited protrusion formation and cell motility, and then reduced the migration of triple-negative breast cancer cells, through increasing the activity of Cdc42 by non-canonical Notch pathway. Phosphorylation of AKT on S473 was surprisingly increased when Notch signaling was inhibited by DAPT. Inhibition of PI3K and AKT by LY294002 and MK2206, respectively, or knockdown of AKT expression by siRNA blocked DAPT-induced activation of Cdc42. Moreover, immunofluorescence staining further showed that DAPT treatment reduced the formation of lamellipodia and induced actin cytoskeleton remodeling. Taken together, these results indicated that DAPT inhibited Notch signaling and consequently activated PI3K/AKT/Cdc42 signaling by non-canonical pathway, facilitated the formation of filopodia and inhibited the assembly of lamellipodia, and finally resulted in the decrease of migration activity of breast cancer cells.
Insights
DAPT, a Notch inhibitor, reduces breast cancer cell migration by activating Cdc42 via a non-canonical pathway. This involves the PI3K/AKT pathway, affecting actin cytoskeleton and cell protrusion formation.
Area of Science:
- Cell Biology
- Cancer Research
- Molecular Biology
Background:
- Tumor cell migration is crucial for cancer metastasis.
- The Notch pathway, when over-activated, promotes cancer cell migration, particularly in breast cancer.
- The precise mechanisms of non-canonical Notch signaling in cell migration remain unclear.
Purpose of the Study:
- To elucidate the non-canonical Notch signaling pathway's role in modulating breast cancer cell migration.
- To investigate the effects of DAPT, a gamma-secretase inhibitor, on triple-negative breast cancer cell motility.
- To identify key molecular players involved in DAPT-mediated inhibition of cancer cell migration.
Main Methods:
- Utilized DAPT (gamma-secretase inhibitor) to study Notch signaling inhibition.
- Assessed cell protrusion formation, cell motility, and migration using various assays.
- Employed PI3K and AKT inhibitors (LY294002, MK2206) and siRNA for AKT knockdown.
- Performed immunofluorescence staining to analyze actin cytoskeleton remodeling and cell morphology.
Main Results:
- DAPT inhibited protrusion formation and cell motility, reducing triple-negative breast cancer cell migration.
- DAPT treatment increased AKT phosphorylation at S473, indicating PI3K/AKT pathway activation.
- Inhibition of PI3K/AKT or AKT knockdown blocked DAPT-induced Cdc42 activation.
- DAPT reduced lamellipodia formation and induced actin cytoskeleton remodeling, favoring filopodia formation.
Conclusions:
- DAPT inhibits Notch signaling, leading to non-canonical activation of the PI3K/AKT/Cdc42 pathway.
- This pathway modulation facilitates filopodia formation while inhibiting lamellipodia assembly.
- Ultimately, DAPT decreases breast cancer cell migration activity through these molecular and cellular changes.
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