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Bringing Light to Transcription: The Optogenetics Repertoire
摘要： The ability to manipulate expression of exogenous genes in particular regions of living organisms has profoundly transformed the way we study biomolecular processes involved in both normal development and disease. Unfortunately, most of the classical inducible systems lack fine spatial and temporal accuracy, thereby limiting the study of molecular events that strongly depend on time, duration of activation, or cellular localization. By exploiting genetically engineered photo sensing proteins that respond to specific wavelengths, we can now provide acute control of numerous molecular activities with unprecedented precision. In this review, we present a comprehensive breakdown of all of the current optogenetic systems adapted to regulate gene expression in both unicellular and multicellular organisms. We focus on the advantages and disadvantages of these different tools and discuss current and future challenges in the successful translation to more complex organisms.
关键词： transcription,optogenetics,cryptochrome,phytochrome B,LOV,gene expression,UVR8,light
更新于2025-09-09 09:28:46
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Impacts of Cys392, Asp393, and ATP on the FAD Binding, Photoreduction, and the Stability of the Radical State of Chlamydomonas reinhardtii Cryptochrome
摘要： Plant cryptochromes (CRYs) are blue-light receptors that regulate the light-dependent growth, development and circadian rhythm. A flavin adenine dinucleotide (FAD) cofactor is bound to the photolyase homology region (PHR) of plant CRYs that can be photoreduced to neutral radical state under blue light. This photoreaction may trigger subsequent signal transduction. Plant CRYs can also bind an ATP adjacent to FAD in a pocket of PHR. Chlamydomonas reinhardtii contains a single plant CRY, named Chlamydomonas photolyase homologue 1 (CPH1). In CPH1, Cys392 and Asp393 is located near the FAD cofactor. Here we showed that replacing Cys392 with Ser had little effect on the properties of CPH1. But the C392N mutant showed a faster photoreduction rate, and a significantly lower oxidation rate of the neutral radical state compared with those of wild type CPH1. Substituting an Asn for Asp393 in CPH1 improved the binding affinity for FAD as well as the stability of the neutral radical; but photoreduction of the mutant was severely inhibited. In the presence of ATP, CPH1 and its mutants exhibited significantly higher binding affinity for FAD and slower oxidation of the neutral radical. These results reveal that the residues at site 392 and the presence of ATP can tune the stability of the neutral radical; the Asp at site 393 is crucial for photoreduction; and the photoreduction rate was not merely determined by the stability of the neutral radical in CPH1.
关键词： flavin radicals,biophysics,photochemistry,cryptochrome,photolyase
更新于2025-09-04 15:30:14