title: Protein Inactivation by Optogenetic Trapping in Living Cells.
abstract: Optogenetic modules that use genetically encoded elements to control protein function in response to light allow for precise spatiotemporal modulation of signaling pathways. As one of optical approaches, LARIAT (Light-Activated Reversible Inhibition by Assembled Trap) is a unique light-inducible inhibition system that reversibly sequesters target proteins into clusters, generated by multimeric proteins and a blue light-induced heterodimerization module. Here we present a method based on LARIAT for optical inhibition of targets in living mammalian cells. In the protocol, we focus on the inhibition of proteins that modulate cytoskeleton and cell cycle, and describe how to transfect, conduct a photo-stimulation, and analyze the data.
authors.affiliation: Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Republic of Korea.
authors.forename: Hyerim
authors.initials: H
authors.lastname: Park
authors.affiliation: Center for Cognition and Sociality, Institute for Basic Science (IBS), Daejeon, Republic of Korea.
authors.forename: Sangkyu
authors.initials: S
authors.lastname: Lee
authors.affiliation: Center for Cognition and Sociality, Institute for Basic Science (IBS), Seoul, 136-791, Republic of Korea. wondo@kaist.ac.kr.|Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon, 305-701, Republic of Korea. wondo@kaist.ac.kr.
authors.forename: Won Do
authors.initials: WD
authors.lastname: Heo
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002455:Cell Division; D056931:Cryptochromes / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods; D041681:NIH 3T3 Cells; D062308:Optogenetics / Q000379:methods*; D051567:Proto-Oncogene Proteins c-vav / Q000235:genetics / Q000378:metabolism; D014162:Transfection / Q000379:methods
chemical_list: D000199:Actins; C109198:CIB1 protein, human; C538966:CRY2 protein, human; D002135:Calcium-Binding Proteins; D056931:Cryptochromes; D051567:Proto-Oncogene Proteins c-vav; C093651:VAV2 protein, human; C490565:Vav2 protein, mouse
keywords: CIB1; Cell division; Clustering; Cryptochrome 2 (CRY2); Cytoskeleton; LARIAT; Optogenetics; Protein inhibition