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Specific contributions of the small GTPases Rho, Rac, and Cdc42 to Dbl transformation
1Department of Molecular Medicine, Veterinary Medical Center, Cornell University, Ithaca, New York 14853, USA.
Abstract:
Dbl is a representative prototype of a growing family of oncogene products that contain the Dbl homology/pleckstrin homology elements in their primary structures and are associated with a variety of neoplastic pathologies. Members of the Dbl family have been shown to function as physiological activators (guanine nucleotide exchange factors) of the Rho-like small GTPases. Although the expression of GTPase-defective versions of Rho proteins has been shown to induce a transformed phenotype under different conditions, their transformation capacity has been typically weak and incomplete relative to that exhibited by dbl-like oncogenes. Moreover, in some cases (e.g. NIH3T3 fibroblasts), expression of GTPase-defective Cdc42 results in growth inhibition. Thus, in attempting to reconstitute dbl-induced transformation of NIH3T3 fibroblasts, we have generated spontaneously activated ("fast-cycling") mutants of Cdc42, Rac1, and RhoA that mimic the functional effects of activation by the Dbl oncoprotein. When stably expressed in NIH3T3 cells, all three mutants caused the loss of serum dependence and showed increased saturation density. Furthermore, all three stable cell lines were tumorigenic when injected into nude mice. Our data demonstrate that all three Dbl targets need to be activated to promote the full complement of Dbl effects. More importantly, activation of each of these GTP-binding proteins contributes to a different and distinct facet of cellular transformation.
Insights
Oncogene Dbl activates Rho GTPases, crucial for cell transformation. Researchers created "fast-cycling" mutants of Cdc42, Rac1, and RhoA, which mimic Dbl
Area of Science:
- Oncogenes and signal transduction pathways.
- Cellular transformation and tumorigenesis.
- Small GTPase regulation and function.
Background:
- Dbl oncoproteins activate Rho-like GTPases, implicated in neoplastic pathologies.
- GTPase-defective Rho proteins show weak or inhibitory transformation capacity.
- Mimicking Dbl activation is key to understanding oncogenesis.
Purpose of the Study:
- To generate spontaneously activated Rho GTPase mutants (Cdc42, Rac1, RhoA).
- To investigate if these mutants can reconstitute Dbl-induced NIH3T3 fibroblast transformation.
- To determine the distinct contributions of each activated GTPase to transformation.
Main Methods:
- Generation of
- fast-cycling
- mutants of Cdc42, Rac1, and RhoA.
- Stable expression of these mutants in NIH3T3 fibroblasts.
- Assessment of cellular transformation phenotypes (serum dependence, saturation density) and tumorigenicity in nude mice.
Main Results:
- All three spontaneously activated mutants induced loss of serum dependence and increased saturation density in NIH3T3 cells.
- Stable cell lines expressing these mutants were tumorigenic upon injection into nude mice.
- Activation of each Dbl target GTPase contributes distinct facets to cellular transformation.
Conclusions:
- Spontaneously activated Cdc42, Rac1, and RhoA mutants effectively mimic Dbl oncoprotein effects.
- Full Dbl-induced transformation requires activation of all three targets.
- Each activated GTP-binding protein plays a unique role in driving cellular transformation and tumorigenesis.