Neuronal Calcium Recording with an Engineered TEV Protease
journal contributionposted on 27.04.2018, 00:00 by Brianna K. O’Neill, Scott T. Laughlin
Technologies for measuring the transient Ca2+ spikes that accompany neural signaling have revolutionized our understanding of the brain. Nevertheless, microscopic visualization of Ca2+ spikes on the time scale of neural activity across large brain regions or in thick specimens remains a significant challenge. The recent development of stable integrators of Ca2+, instead of transient reporters, provides an avenue to investigate neural signaling in otherwise challenging systems. Here, we describe an engineered Ca2+-sensing enzyme consisting of a split Tobacco Etch Virus (TEV) protease with each half tethered to a calmodulin or M13 Ca2+ binding domain. This Split TEV, Ca2+ Activated Neuron Recorder (SCANR) remains separate and catalytically incompetent until a spike in cellular Ca2+ triggers its reconstitution and the subsequent turnover of a caged, genetically encoded reporter substrate. We report the identification of a successful Ca2+-sensing split TEV from a library of chimeras and deployment of the enzyme in primary rat hippocampal neurons.