Genetically encoded photoswitching of actin assembly through the Cdc42-WASP-Arp2/3 complex pathway.
General methods to engineer genetically encoded, reversible, light-mediated control over protein function would be useful in many areas of biomedical research and technology. We describe a system that yields such photo-control over actin assembly. We fused the Rho family GTPase Cdc42 in its GDP-bound form to the photosensory domain of phytochrome B (PhyB) and fused the Cdc42 effector, the Wiskott-Aldrich Syndrome Protein (WASP), to the light-dependent PhyB-binding domain of phytochrome interacting factor 3 (Pif3). Upon red light illumination, the fusion proteins bind each other, activating WASP, and consequently stimulating actin assembly by the WASP target, the Arp2/3 complex. Binding and WASP activation are reversed by far-red illumination. Our approach, in which the biochemical specificity of the nucleotide switch in Cdc42 is overridden by the light-dependent PhyB-Pif3 interaction, should be generally applicable to other GTPase-effector pairs.
Leung
Daisy W
DW
Howard Hughes Medical Institute, Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
Otomo
Chinatsu
C
Chory
Joanne
J
Rosen
Michael K
MK
D051376:Actin-Related Protein 2-3 Complex / Q000378:metabolism*; D000199:Actins / Q000378:metabolism*; D017360:Arabidopsis / Q000528:radiation effects; D029681:Arabidopsis Proteins / Q000378:metabolism; D051792:Basic Helix-Loop-Helix Proteins / Q000378:metabolism; D005818:Genetic Engineering; D006153:Guanosine Diphosphate / Q000378:metabolism; D008027:Light; D055537:Light Signal Transduction / Q000528:radiation effects; D051939:Phytochrome B / Q000378:metabolism; D011485:Protein Binding; D051316:Wiskott-Aldrich Syndrome Protein / Q000378:metabolism*; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism*
D051376:Actin-Related Protein 2-3 Complex; D000199:Actins; D029681:Arabidopsis Proteins; D051792:Basic Helix-Loop-Helix Proteins; D006153:Guanosine Diphosphate; D051939:Phytochrome B; D051316:Wiskott-Aldrich Syndrome Protein; D020764:cdc42 GTP-Binding Protein; C116162:PIF3 protein, Arabidopsis