Lutz Anne P, Renicke Christian, Taxis Christof
Department of Biology/Genetics, Philipps-University Marburg, Karl-von-Frisch-Str. 8, 35043, Marburg, Germany.
Methods Mol Biol. 2016;1408:67-78. doi: 10.1007/978-1-4939-3512-3_5.
Regulation of protein stability is a fundamental process in eukaryotic cells and pivotal to, e.g., cell cycle progression, faithful chromosome segregation, or protein quality control. Synthetic regulation of protein stability requires conditional degradation sequences (degrons) that induce a stability switch upon a specific signal. Fusion to a selected target protein permits to influence virtually every process in a cell. Light as signal is advantageous due to its precise applicability in time, space, quality, and quantity. Light control of protein stability was achieved by fusing the LOV2 photoreceptor domain of Arabidopsis thaliana phototropin1 with a synthetic degron (cODC1) derived from the carboxy-terminal degron of ornithine decarboxylase to obtain the photosensitive degron (psd) module. The psd module can be attached to the carboxy terminus of target proteins that are localized to the cytosol or nucleus to obtain light control over their stability. Blue light induces structural changes in the LOV2 domain, which in turn lead to activation of the degron and thus proteasomal degradation of the whole fusion protein. Variants of the psd module with diverse characteristics are useful to fine-tune the stability of a selected target at permissive (darkness) and restrictive conditions (blue light).
蛋白质稳定性的调控是真核细胞中的一个基本过程，对于细胞周期进程、准确的染色体分离或蛋白质质量控制等至关重要。蛋白质稳定性的合成调控需要条件性降解序列（降解子），该序列在特定信号作用下引发稳定性开关。与选定的靶蛋白融合可影响细胞内几乎所有过程。光作为信号具有优势，因为它在时间、空间、质量和数量上具有精确的适用性。通过将拟南芥向光素1的LOV2光感受器结构域与源自鸟氨酸脱羧酶羧基末端降解子的合成降解子（cODC1）融合，实现了对蛋白质稳定性的光控，从而获得光敏降解子（psd）模块。psd模块可连接到定位于细胞质或细胞核的靶蛋白的羧基末端，以实现对其稳定性的光控。蓝光诱导LOV2结构域发生结构变化，进而导致降解子激活，从而使整个融合蛋白被蛋白酶体降解。具有不同特性的psd模块变体有助于在允许条件（黑暗）和限制条件（蓝光）下微调选定靶标的稳定性。
