FRET Imaging and cis-regulatory Plasticity of Shp2 Conformation

Signaling molecules and their activities are well coordinated in space and time to regulate cellular functions in response to mechanical and chemical microenvironment. Based on fluorescent resonance energy transfer (FRET), we have developed genetically encoded biosensors to monitor the dynamic molecular activities (such as Src and FAK activities) in live cells at subcellular compartments when cells interact with their neighbors or mechanical/chemical microenvironment. In a recent study, we show that a ubiquitous signaling protein, Src Homology 2 (SH2) Domain-containing Protein Tyrosine Phosphatase 2 (Shp2), displayed unexpected plasticity of conformational changes via intramolecular interactions within Shp2 (cis-interaction). Utilizing Shp2 biosensors based on fluorescence resonance energy transfer (FRET), we found that two phosphorylated regulatory tyrosines upon stimulated phosphorylation can compete for the cis-interaction of the same SH2 domain within Shp2 to achieve plasticity. The antagonistic combination of contextual amino acid sequence and position (e.g. favorable position combined with adverse sequence) can create a relatively small difference between the two phosphorylated tyrosines in their overall competitiveness for cis-interaction. Enlarging this difference by swapping the sequences at the two tyrosine positions resulted in loss of conformational plasticity and reprogrammed downstream ERK signaling dynamics. Thus, while the combinatorial effect of specific sequence and position of DNA cis-regulatory elements on tuning gene expression has been well studied, our results unraveled a new and simple strategy to achieve cis-regulatory plasticity of protein conformation by coordinating the combination of sequence and position. We suggest that this strategy can serve as a general and basic design principle for natural and synthetic proteins, with their conformations and functions tunable by cis-interaction to regulate downstream physiological consequences. These proteins with plasticity can serve as programmable building blocks or nodes for higher order molecular machines and networks.