Bringing Light to Transcription: The Optogenetics Repertoire

Abstract

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.

Keywords: LOV; UVR8; cryptochrome; gene expression; light; optogenetics; phytochrome B; transcription.

FIGURE 1

Schematic layout of red-light inducible system (PhyB-PIF). In presence of the chromophore and upon red light irradiation (∼630 nm), PhyB interacts with its binding partner PIF to reconstitute a split transcription factor (DBD and AD) and trigger gene expression. Conversely, under far-red light irradiation (>720 nm), the PhyB-PIF complex dissociates and transcription ceases. The yellow box represents the exogenous chromophore. Arrows depict direction of activity and light conditions: the red arrow represents red light-dependent activation (∼630 nm), the dark red arrow represents far-red light-dependent inactivation (>720 nm), solid black arrow represents interaction of PhyB and chromophore independent of light.

FIGURE 2

Schematic illustration of blue light-dependent optogenetic strategies. (A) Upon blue light irradiation (∼450 nm), Cry2 dimerizes with CIBN to induce activation of a split transcription factor (DBD and AD) and promotes gene expression. (B) In the presence of blue light, EL222 undocks HTH-DBD facilitating homodimerization of two EL222 proteins which, then, bind to C120 promoter to trigger transcription. (C) Upon blue light irradiation, a monomeric form of LOV uncages a small NLS causing the system (LANS or LINuS) to accumulate in the nucleus. In the absence of blue light or darkness, a counter-active NES peptide outside the LOV-cage allows for the system to accumulate in the cytoplasm. (D) In darkness, an NLS peptide outside the LOV-cage allows for the optogenetic system (LINX or LEXY) to accumulate in the nucleus and promote gene expression. After blue light irradiation, LOV uncages a small counter-active NES causing the system to translocate to the cytoplasm and transcription ceases. (E) In the absence of light, LINX in combination with a light dependent dimer (iLid) accumulates in the nucleus and triggers gene expression. Upon blue irradiation, LINX-iLid translocates to the cytoplasm where it dimerizes with a second iLid dimer sequestering the system to the mitochondria. (F) In the cytoplasm, LOV dark-state selectively binds to the peptide Zdk sequestering LOV to the mitochondria. Upon light irradiation, LOV’s conformational change allows for the release of the system from zdk and translocate to the nucleus. Arrows depict direction of activity and light conditions: blue arrow (∼450 nm) represents blue light activation, while dashed black arrow represents darkness or absence of light stimuli. For simplicity, light purple boxes inside the photo-sensing protein represent the chromophores.

FIGURE 3

Schematic illustration of UV-dependent system (UVR8-COP1). In darkness, UVR8 accumulates in cytoplasm as homodimers. Upon UV-B light irradiation, UVR8 monomerizes and associates with its binding partner COP1 to reconstitute a split transcription factor (DBD and AD). Then, the complex translocates to the nucleus and promotes gene expression. Arrows depict direction of activity and light conditions: the purple arrow (∼315 nm) represents UV-B light-dependent activation, while the dashed black arrow represents darkness or absence of light stimuli.