Bidirectional approaches for optogenetic regulation of gene expression in mammalian cells using Arabidopsis cryptochrome 2

• Published in: Nucleic acids research Vol. 45; no. 20; p. e167
• Main Authors: Pathak, Gopal P, Spiltoir, Jessica I, Höglund, Camilla, Polstein, Lauren R, Heine-Koskinen, Sari, Gersbach, Charles A, Rossi, Jari, Tucker, Chandra L
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 16-11-2017
• Subjects: Animals; Animals, Genetically Modified; Arabidopsis - genetics; Arabidopsis Proteins - genetics; Basic Helix-Loop-Helix Transcription Factors - genetics; Cell Line; Cryptochromes - genetics; DNA-Binding Proteins - genetics; Gene Expression Regulation; HEK293 Cells; Humans; Light; Methods Online; Optogenetics - methods; Transcriptional Activation - genetics; Zebrafish - genetics
• ISSN: 0305-1048; 1362-4962
• DOI: 10.1093/nar/gkx260

Optogenetic tools allow regulation of cellular processes with light, which can be delivered with spatiotemporal resolution. In previous work, we used cryptochrome 2 (CRY2) and CIB1, Arabidopsis proteins that interact upon light illumination, to regulate transcription with light in yeast. While adopting this approach to regulate transcription in mammalian cells, we observed light-dependent redistribution and clearing of CRY2-tethered proteins within the nucleus. The nuclear clearing phenotype was dependent on the presence of a dimerization domain contained within the CRY2-fused transcriptional activators. We used this knowledge to develop two different approaches to regulate cellular protein levels with light: a system using CRY2 and CIB1 to induce protein expression with light through stimulation of transcription, and a system using CRY2 and a LOV-fused degron to simultaneously block transcription and deplete protein levels with light. These tools will allow precise, bi-directional control of gene expression in a variety of cells and model systems.