Pook Bastian, Goenrich Juri, Hasenjäger Sophia, Essen Lars-Oliver, Spadaccini Roberta, Taxis Christof
Unit for Structural Biochemistry, Department of Chemistry, Philipps-University Marburg, 35032 Marburg, Germany.
Department of Biology/Genetics, Philipps-University Marburg, 35043 Marburg, Germany.
ACS Synth Biol. 2021 Dec 17;10(12):3411-3421. doi: 10.1021/acssynbio.1c00350. Epub 2021 Nov 19.
Optogenetic tools have been proven to be useful in regulating cellular processes via an external signal. Light can be applied with high spatial and temporal precision as well as easily modulated in quantity and quality. Natural photoreceptors of the light oxygen voltage (LOV) domain family have been characterized in depth, especially the LOV2 domain of (As) phototropin 1 and its derivatives. Information on the behavior of LOV2 variants with changes in the photocycle or the light response has been recorded. Here, we applied well-described photocycle mutations on the AsLOV2 domain of a photosensitive transcription factor (psTF) as well as its variant that is part of the photosensitive degron (psd) psd3 in . and measurements revealed that each photoreceptor component of the light-sensitive transcription factor and the psd3 module can be modulated in its light sensitivity by mutations that are known to prolong or shorten the dark-reversion time of AsLOV2. Yet, only two of the mutations showed differences in the behavior in the context of the psd3 module. For the AsLOV2 domain in the context of the psTF, we observed different characteristics for all four variants. Molecular dynamics simulations showed distinct influences of the shortened Jα helix and the V416L mutation in the context of the psd3 photoreceptor. In conclusion, we demonstrated the tunability of two optogenetic tools with a set of mutations that affect the photocycle of the inherent photoreceptors. As these optogenetic tools are concurrent in their action, pleiotropic effects on target protein abundance are achievable with the simultaneous action of the diverse photoreceptor variants.
光遗传学工具已被证明可通过外部信号调节细胞过程。光可以以高空间和时间精度施加，并且在数量和质量上易于调节。光氧电压（LOV）结构域家族的天然光感受器已得到深入表征，尤其是（As）向光素1的LOV2结构域及其衍生物。已记录了关于LOV2变体在光循环或光响应变化时的行为信息。在这里，我们对光敏转录因子（psTF）的AsLOV2结构域及其作为光敏降解子（psd）psd3一部分的变体应用了描述清晰的光循环突变。 和 测量结果表明，光敏转录因子和psd3模块的每个光感受器组件的光敏感性都可以通过已知能延长或缩短AsLOV2暗恢复时间的突变进行调节。然而，在psd3模块的背景下，只有两个突变在 行为上表现出差异。对于psTF背景下的AsLOV2结构域，我们观察到所有四个变体都有不同的特征。分子动力学模拟显示，在psd3光感受器的背景下，缩短的Jα螺旋和V416L突变有明显影响。总之，我们用一组影响固有光感受器光循环的突变证明了两种光遗传学工具的可调性。由于这些光遗传学工具在作用上是并行的，不同光感受器变体的同时作用可实现对靶蛋白丰度的多效性影响。
