Related Experiment Video
Updated: Jul 11, 2026
09:58
RhoC GTPase Activation Assay
Published on: August 22, 2010
The RhoGEF domain of p210 Bcr-Abl activates RhoA and is required for transformation
S Sahay1, N L Pannucci, G M Mahon
1Department of Microbiology and Molecular Genetics, New Jersey Medical School-University Hospital Cancer Center, University of Medicine and Dentistry of New Jersey, Newark, NJ 7103, USA.
Oncogene
|October 9, 2007
Summary
The Rho-specific guanine nucleotide exchange factor (RhoGEF) domain of the BCR-ABL fusion protein is constitutively active in cancer. This RhoGEF activity is crucial for BCR-ABL
Area of Science:
- Oncogenic signaling pathways
- Molecular mechanisms of cancer
- Protein-protein interactions in cancer
Background:
- The BCR-ABL oncogene is a fusion protein central to chronic myeloid leukemia.
- BCR-ABL includes a Rho-specific guanine nucleotide exchange factor (RhoGEF) domain from BCR.
- This RhoGEF domain is typically autoinhibited in BCR but constitutively active in BCR-ABL.
Purpose of the Study:
- To investigate the role of the RhoGEF domain's constitutive activation in p210 BCR-ABL.
- To determine if RhoGEF activity is essential for BCR-ABL's transforming capabilities.
- To delineate which aspects of BCR-ABL-mediated transformation depend on RhoGEF function.
Main Methods:
- Site-directed mutagenesis of the RhoGEF domain in p210 BCR-ABL.
- Assays for RhoA activation.
- Assessment of tyrosine kinase activity.
- Analysis of anchorage-independent growth and growth factor independence in myeloid cells.
Main Results:
- Mutations disabling RhoGEF activity inhibited RhoA activation without affecting tyrosine kinase activity.
- The RhoGEF mutant of p210 BCR-Abl showed impaired anchorage-independent growth.
- Despite impaired transformation, the RhoGEF mutant retained the ability to confer growth factor independence.
Conclusions:
- The RhoGEF domain of p210 BCR-ABL possesses a gain-of-function activity.
- Catalytically active RhoGEF is required for specific transforming activities of BCR-ABL, such as anchorage-independent growth.
- Not all oncogenic functions of BCR-ABL are dependent on its RhoGEF activity.
More Related Videos
11:28Affinity Precipitation of Active Rho-GEFs Using a GST-tagged Mutant Rho Protein (GST-RhoA(G17A)) from Epithelial Cell Lysates
Published on: March 31, 2012
10:27Spatio-Temporal Manipulation of Small GTPase Activity at Subcellular Level and on Timescale of Seconds in Living Cells
Published on: March 9, 2012
Related Concept Videos
01:24Small GTPases - Ras and Rho
Ras and Rho are small monomeric GTPases that act downstream of receptor tyrosine kinase (RTK) and regulate various cellular processes. These GTPases switch between active and inactive states by binding to guanine nucleotides.
Three regulatory proteins control their activity:
Three regulatory proteins control their activity:
02:38The Ras Gene
The Ras-gene-encoded proteins are regulators of signaling pathways controlling cell proliferation, differentiation, or cell survival. The Ras-gene family in humans constitutes three primary members—the HRas, NRas, and KRas. These genes code for four functionally distinct yet closely related proteins—the HRas, NRas, KRas4A, and KRas4B. The involvement of mutant Ras genes in human cancer was first discovered in 1982 and is among the most common causes of human tumorigenesis.
Ras is a superfamily...
Ras is a superfamily...
01:21Cell Polarization by Rho Proteins
Cell polarity is the asymmetric distribution of cellular and membrane components, making one side of the cell different from the other. This polarity is essential to many processes such as embryogenesis, axon migration, glucose transport across epithelial cells, and directional cell migration. A migrating cell responds to intracellular or extracellular signals via molecular cascades that reorganize the actin cytoskeleton to establish this polarity. In these cells, the Rho family proteins Cdc42,...
01:07MAPK Signaling Cascades
Mitogen-activated protein kinase, or MAPK pathway, activates three sequential kinases to regulate cellular responses such as proliferation, differentiation, survival, and apoptosis. The canonical MAPK pathway starts with a mitogen or growth factor binding to an RTK. The activated RTKs stimulate Ras, which recruits Raf or MAP3 Kinase (MAPKKK), the first kinase of the MAPK signaling cascade. Raf further phosphorylates and activates MEK or MAP2 Kinases (MAPKK), which in turn phosphorylates MAP...
01:03Rous Sarcoma Virus (RSV) and Cancer
Rous Sarcoma virus or RSV was discovered by F. Peyton Rous in the year 1911 as a filterable transmissible agent that could cause tumors in chickens. He won a Nobel Prize for this discovery in 1966. His experiments clearly demonstrated that some cancers could be caused by infectious agents and led to the discovery of many more cancer-causing viruses in animals as well as humans.
RSV is a retrovirus that contains two copies of a plus-strand RNA genome. Its genome consists of four main open...
RSV is a retrovirus that contains two copies of a plus-strand RNA genome. Its genome consists of four main open...
01:14Rab Proteins
Rab proteins constitute the largest family of monomeric GTPases, of which 70 members are present in humans. Rab proteins and their effectors regulate consecutive stages of vesicle transport such as vesicle transport, docking, and fusion to the correct recipient membrane.
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...