Induced Recruitment & Activation of Mammalian Rho GTPase

Research Summary

The iDimerize Inducible Heterodimerization technology is used to understand the role of Rho GTPase Cdc42 in regulating actin polymerization. Authors show that induced recruitment of Cdc42 to the membrane is sufficient for producing actin-based membrane protrusions. Paper: Castellano et al. (1999) J. Cell Sci. 113:2955–61.

Ligand induced dimerization & activation of Cdc42 in mammalian cells

Advantages of inducible dimerization

Experimental design

An inducible system for recruiting Cdc42 to the plasma membrane was created in rat basophilic leukemia (RBL-2H3) cells. Two copies of the DmrA domain were anchored to the cell membrane using a signal sequence. The cytosolic, constitutively active form of Cdc42 was expressed fused to a DmrC domain, allowing inducible translocation to the plasma membrane upon addition of A/C Heterodimerizer ligand to growth media.*

Key Results

  1. Localized recruitment of Cdc42 to the membrane was sufficient for producing the signature phenotype of actin-based membrane protrusions.

  2. A parallel study showed that membrane recruitment of Rac1 triggers actin-dependent particle internalization via phagocytosis (Castellano, F. 2000).

*

Studies are presented using iDimerize components. For complete experimental details please refer to the original publication. The optimal position and copy number of fusion domains varies per study and must be determined experimentally.

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