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Article Abstract
GCaMP, one popular type of genetically-encoded Ca indicator, has been associated with various side-effects. Here we unveil the intrinsic problem prevailing over different versions and applications, showing that GCaMP containing CaM (calmodulin) interferes with both gating and signaling of L-type calcium channels (Ca1). GCaMP acts as an impaired apoCaM and Ca/CaM, both critical to Ca1, which disrupts Ca dynamics and gene expression. We then design and implement GCaMP-X, by incorporating an extra apoCaM-binding motif, effectively protecting Ca1-dependent excitation-transcription coupling from perturbations. GCaMP-X resolves the problems of detrimental nuclear accumulation, acute and chronic Ca dysregulation, and aberrant transcription signaling and cell morphogenesis, while still demonstrating excellent Ca-sensing characteristics partly inherited from GCaMP. In summary, CaM/Ca1 gating and signaling mechanisms are elucidated for GCaMP side-effects, while allowing the development of GCaMP-X to appropriately monitor cytosolic, submembrane or nuclear Ca, which is also expected to guide the future design of CaM-based molecular tools.
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| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5904127 | PMC |
| http://dx.doi.org/10.1038/s41467-018-03719-6 | DOI Listing |
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