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Piatkevich, Kiryl D; Jung, Erica E; Straub, Christoph; Linghu, Changyang; Park, Demian; Suk, Ho-Jun; Hochbaum, Daniel R; Goodwin, Daniel; Pnevmatikakis, Eftychios; Pak, Nikita; Kawashima, Takashi; Yang, Chao-Tsung; Rhoades, Jeffrey L; Shemesh, Or; Asano, Shoh; Yoon, Young-Gyu; Freifeld, Limor; Saulnier, Jessica L; Riegler, Clemens; Engert, Florian; Hughes, Thom; Drobizhev, Mikhail; Szabo, Balint; Ahrens, Misha B; Flavell, Steven W; Sabatini, Bernardo L; Boyden, Edward S
Nature chemical biology, 04/2018, Letnik: 14, Številka: 4Journal Article
We developed a new way to engineer complex proteins toward multidimensional specifications using a simple, yet scalable, directed evolution strategy. By robotically picking mammalian cells that were identified, under a microscope, as expressing proteins that simultaneously exhibit several specific properties, we can screen hundreds of thousands of proteins in a library in just a few hours, evaluating each along multiple performance axes. To demonstrate the power of this approach, we created a genetically encoded fluorescent voltage indicator, simultaneously optimizing its brightness and membrane localization using our microscopy-guided cell-picking strategy. We produced the high-performance opsin-based fluorescent voltage reporter Archon1 and demonstrated its utility by imaging spiking and millivolt-scale subthreshold and synaptic activity in acute mouse brain slices and in larval zebrafish in vivo. We also measured postsynaptic responses downstream of optogenetically controlled neurons in C. elegans.
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