Year: 2026
Type: preprint
Abstract: Calcium-calmodulin (CaM)-dependent protein kinase II (CaMKII) is a key mediator of complex physiological processes throughout the body, from the brain to the reproductive system, where CaMKII translates spatiotemporally dynamic calcium elevations into specific biological functions. Directly visualizing CaMKII activity dynamics in living cells using genetically encoded fluorescent biosensors can thus provide crucial insights into the molecular regulation of health and disease. Yet the ability to sensitively and specifically monitor endogenous CaMKII activity in physiologically relevant contexts is limited by the lack of sensors that can achieve robust, quantitative visualization of CaMKII responses. Here, we leveraged a recent serine/threonine kinome-wide substrate atlas to rationally engineer a powerful suite of Förster resonance energy transfer (FRET)-based CaMKII kinase activity reporters with high specificity, sensitivity, and signal-to-noise ratio. Using these biosensors, we were able to sensitively and robustly visualize endogenous CaMKII activity dynamics in both cultured cell lines and primary cells, including cardiomyocytes, oocytes, and neurons. We further utilized 2pFLIM imaging of organotypic hippocampal slices to quantitatively track LTP-induced CaMKII activity within single dendritic spines, highlighting a major advance in the study of physiological CaMKII signaling.
Authors: Sohum Mehta, Nidhi A. Thaker, Kengo Adachi, Christopher Y. Ko, Bian Liu, Sai S. Divkaruni, Oguz C. Koc, Anne C. Lyons, Samantha A. Sanchez, Pauline Löffler, Yoshihisa Nakahata, Jody L. Martin, Jared L. Johnson, Tomer M. Yaron-Barir, Lewis C. Cantley, Martin J. Lohse, Andreas Bock, Donald M. Bers, Ryohei Yasuda, Rafael Fissore, Richard L. Huganir, Margaret M. Stratton, Jin Zhang
Institutions: University of California San Diego, University of Massachusetts Amherst, Max Planck Florida Institute for Neuroscience, University of California, Davis, Johns Hopkins University, Johns Hopkins Medicine, Leipzig University, Harvard University, Dana-Farber Cancer Institute, Boston Children's Hospital, University Medical Center of the Johannes Gutenberg University Mainz, Moores Cancer Center
Language: English
Subfield: Biophysics
Domain: Life Sciences
Open Access status: green
Funders: Oregon Health and Science University, National Institutes of Health, University of California, Davis, University of California, San Diego, National Heart, Lung, and Blood Institute, National Institute of Mental Health, National Cancer Institute, National Institute of General Medical Sciences, National Institute of Neurological Disorders and Stroke, National Institute of Biomedical Imaging and Bioengineering, Eunice Kennedy Shriver National Institute of Child Health and Human Development
Awards: Regulation of Ca2+ influx in mouse oocytes and eggs during maturation and fertilization to improve assisted reproductive technologies and modulate fertility, Development of kinase biosensors for multiplex neuronal imaging of signaling pathways in behaving mice, Illuminating ER Lumen AMPK Dynamics for Maintaining Vascular Homeostasis via ACE2, Mechanisms of Ras signaling in single synapses, Mammalian Fertilization: Identifying the second sperm factor that induces residual calcium oscillations and its contributions to egg activation, Role of Phosphoinositides and Protein Kinases in the control of Cancer Metabolism, Molecular Pathogenesis of the Hamartoma Syndromes, Neuronal Intracellular Signaling Underlying Synaptic, Circuit and Behavioral Plasticity, Ultrasensitive kinase biosensors for multiplex imaging of coordinated spatiotemporal signaling in cancer-immune interactions, Genetics and Biology of Pancreatic Ductal Adenocarcinoma, Unraveling the molecular events driven by CaMKII in Ca2+-coupled cells, Training in Multi-scale Analysis of Biological Structure and Function, Live-cell Activity Architecture in Cancer, Multiplex Imaging of Brain Activity and Plasticity with Optimized FRET/FLIM-based Sensors, Calmodulin and Regulation of Cardiac Ryanodine Receptor and CaMKII, CaMKII activation and regulation in adult cardiac myocytes, Biotechnology Training Program in Applied Life Sciences
OpenAlex ID: W7162068734