A covalently linked probe to monitor local membrane properties surrounding plasma membrane proteins

We present a method probing the local membrane environment surrounding membrane proteins in the plasma membrane by linking a solvatochromic dye to the target, and provide evidence for lipid-based sorting. The insulin receptor environment changed upon insulin stimulation revealing dynamic local changes and distance dependence. Functional membrane proteins in the plasma membrane are suggested to have specific membrane environments that play important roles to maintain and regulate their function. However, the local membrane environments of membrane proteins remain largely unexplored due to the lack of available techniques. We have developed a method to probe the local membrane environment surrounding membrane proteins in the plasma membrane by covalently tethering a solvatochromic, environment-sensitive dye, Nile Red, to a GPI-anchored protein and the insulin receptor through a flexible linker. The fluidity of the membrane environment of the GPI-anchored protein depended upon the saturation of the acyl chains of the lipid anchor. The local environment of the insulin receptor was distinct from the average plasma membrane fluidity and was quite dynamic and heterogeneous. Upon addition of insulin, the local membrane environment surrounding the receptor specifically increased in fluidity in an insulin receptor-kinase dependent manner and on the distance between the dye and the receptor.

Automated summary

This study presents a new method to probe the local membrane environment surrounding membrane proteins and provides evidence for lipid-based sorting. The environment surrounding the insulin receptor changed dynamically upon insulin stimulation, revealing its dependence on distance. Functional membrane proteins in the plasma membrane have specific membrane environments that are crucial for their maintenance and regulation.