Repurposing protein degradation for optogenetic modulation of protein activities

Abstract

Non-neuronal optogenetic approaches empower precise regulation of protein dynamics in live cells but often require target-specific protein engineering. To address this challenge, we developed a generalizable light modulated protein stabilization system (GLIMPSe) to control intracellular protein level independent of its functionality. We applied GLIMPSe to control two distinct classes of proteins: mitogen-activated protein kinase phosphatase 3 (MKP3), a negative regulator of the extracellular signal-regulated kinase (ERK) pathway, as well as a constitutively active form of MEK (CA MEK), a positive regulator of the same pathway. Kinetics study showed that light-induced protein stabilization could be achieved within 1 minute of blue light stimulation.GLIMPSe enables target-independent optogenetic control of protein activities and therefore minimizes the systematic variation embedded within different photoactivatable proteins. Overall, GLIMPSe promises to achieve light-mediated post-translational stabilization of a wide array of target proteins in live cells.

Targeted Protein Degradation through Fast Optogenetic Activation and Its Application to the Control of Cell Signaling

“…16 Examples include a photocaged small molecule dimerizer of the auxin-inducible domain (AID) and optogenetic approaches, such as LOV2-degron fusions for optical activation of protein degradation (B-LID) 17−19 and photoactivated cleavage of appended degron domain for optical deactivation of degradation (GLIMPSe). 20 Optical control over PROTAC function has been achieved through synthetic analogs bearing photoswitchable azobenzene linkers 21,22 and light-removable caging groups. 23−25 While the technologies mentioned above offer optical control of target protein activity, these designs include certain limitations.…”

Engineering Photosensory Modules of Non-Opsin-Based Optogenetic Actuators

“…Although the development of these system required substantial efforts in protein engineering, this versatile strategy enables the light-dependent protein translocation in a simpler way by fusing the biofunctional domain to the LINuS or LEXY. In addition to optical control of gene expression by manipulating the localization of transcription factor [ 20 , 21 ], LINuS and LEXY systems have also been used in optogenetic regulation of cofilin-1 for controlling F-actin assembly [ 99 ], manipulating of the endogenous p53 protein levels [ 100 ], and optogenetic translocation of TEV protease for manipulating protein degradation [ 101 ].…”

Lights up on organelles: Optogenetic tools to control subcellular structure and organization

“…Evolved LOV contains mutations to improve the photocaging of Tobacco Etch Virus (TEV) protease cleavage sites. GLIMPSe introduces these cleavage sites between the C‐terminus of POI‐eLOV Jα helices and a series of degron signals derived from Xenopus laevis that target the uncleaved fusion for degradation (Mondal et al, 2019). These degron signals can be cleaved off by TEV proteases upon their light‐triggered release from the nucleus, where they are sequestered by an NLS fused to LEXY.…”