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Cdc42 and Ras cooperate to mediate cellular transformation by intersectin-L
Jian-Bin Wang1, Wen Jin Wu, Richard A Cerione
1Department of Molecular Medicine, Veterinary Medical Center, Cornell University, Ithaca, New York 14853, USA.
The Journal of Biological Chemistry
|April 13, 2005
Summary
The study reveals that a specific intersectin-L protein fragment (SH3A-C2) transforms cells by cooperating with Ras, not by altering epidermal growth factor receptor (EGFR) signaling. This finding clarifies mechanisms of cellular transformation involving Cdc42.
Area of Science:
- Cell Biology
- Molecular Biology
- Oncology
Background:
- Cdc42 (a Ras-related GTP-binding protein) regulates key cellular processes including actin cytoskeleton organization, membrane trafficking, cell-cycle progression, and malignant transformation.
- Previous work demonstrated that a fast-cycling Cdc42 mutant (Cdc42(F28L)) transforms NIH 3T3 cells by disrupting epidermal growth factor receptor (EGFR)-Cbl interactions and subsequent EGFR down-regulation.
Purpose of the Study:
- To investigate the relationship between Cdc42 hyperactivation and its impact on EGFR signaling, leading to cellular transformation.
- To elucidate the role of different intersectin-L protein domains in modulating Cdc42 activity and cellular transformation.
Main Methods:
- Expression of various intersectin-L protein fragments in fibroblasts.
- Assessing guanine nucleotide exchange factor (GEF) activity of intersectin-L fragments on Cdc42.
- Evaluating cellular transformation phenotypes, including growth in low serum and colony formation in soft agar.
- Analyzing effects on EGFR levels and epidermal growth factor (EGF)-coupled signaling through ERK.
Main Results:
- A truncated intersectin-L fragment (SH3A-C2) containing DH, PH, and C2 domains exhibited significant GEF activity, strongly activating Cdc42.
- SH3A-C2 potently stimulated cell growth in low serum and colony formation in soft agar, indicating potent transforming activity.
- Unlike the Cdc42(F28L) mutant, SH3A-C2 did not significantly alter EGFR levels or the duration of EGF-induced ERK signaling.
- SH3A-C2 mediated cellular transformation primarily through cooperation between Ras and Cdc42, rather than through EGFR pathway modulation.
Conclusions:
- The intersectin-L SH3A-C2 fragment transforms fibroblasts through a mechanism involving Ras-Cdc42 cooperation.
- This pathway of cellular transformation is distinct from the EGFR-dependent mechanism observed with the fast-cycling Cdc42(F28L) mutant.
- These findings highlight the diverse roles of intersectin-L in regulating cell signaling and transformation.