1.
Using split protein reassembly strategy to optically control PLD enzymatic activity.
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Yao, Y
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Lou, X
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Jianxu, L
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Zhu, M
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Qian, X
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Liu, J
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Zhang, L
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Zhang, P
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He, L
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Li, H
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Xu, Y
Abstract:
Phospholipase D (PLD) and phosphatidic acid (PA) play a spatio-temporal role in regulating diverse cellular activities. Although current methodologies enable optical control of the subcellular localization of PLD and by which influence local PLD enzyme activity, the overexpression of PLD elevates the basal PLD enzyme activity and further leads to increased PA levels in cells. In this study, we employed a split protein reassembly strategy and optogenetic techniques to modify superPLD (a PLDPMF variant with a high basal activity). We splited this variants into two HKD domains and fused these domains with optogenetic elements and by which we achieved light-mediated dimerization of the two HKD proteins and then restored the PLD enzymatic activity.
2.
Optogenetic strategies for optimizing the performance of biosensors of membrane phospholipids in live cells.
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Yao, Y
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Lou, X
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Du, G
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Jin, L
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Jianxu, L
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Liu, J
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Chen, Y
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Cheng, S
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Zhao, T
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Ke, S
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Zhang, L
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Zhang, P
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Xu, Y
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He, L
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Li, H
Abstract:
High-performance biosensors are crucial for elucidating the spatiotemporal regulatory roles and dynamics of membrane lipids, but there is a lack of improvement strategies for biosensors with low sensitivity and low-content substrates detection. Here we developed universal optogenetic strategies to improve a set of membrane biosensors by trapping them into specific region and further reducing the background signal, or by optically-controlled phase separation for membrane lipids detection and tracking. These improved biosensors were superior to typical tools and light simulation would enhance their detection performance and resolution, which might contribute to the design and optimization of other biosensors.