Simultaneous detection of membrane contact dynamics and associated Ca2+ signals by reversible chemogenetic reporters

Garcia Casas, P.; Rossini, M.; Pavenius, L.; Saeed, M.; Arnst, N.; Sonda, S.; Bruzzone, M.; Berno, V.; Raimondi, A.; Sassano, M. L.; Naia, L.; Agostinis, P.; Sturlese, M.; Niemeyer, B. A.; Brismar, H.; Ankarcrona, M.; Gautier, A.; Pizzo, P.; Filadi, R.

doi:10.1101/2023.12.28.573515

bioRxiv · 10.1101/2023.12.28.573515

Simultaneous detection of membrane contact dynamics and associated Ca2+ signals by reversible chemogenetic reporters

Garcia Casas, P.·Rossini, M.·Pavenius, L.·Saeed, M.·Arnst, N.·Sonda, S.·Bruzzone, M.·Berno, V.·Raimondi, A.·Sassano, M. L.·Naia, L.·Agostinis, P.·Sturlese, M.·Niemeyer, B. A.·Brismar, H.·Ankarcrona, M.·Gautier, A.·Pizzo, P.·Filadi, R.

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Abstract

Membrane contact sites (MCSs) are hubs allowing various cell organelles to coordinate their activities. The dynamic nature of these sites and their small size hinder analysis by current imaging techniques. To overcome these limitations, we here designed a series of reversible chemogenetic reporters incorporating improved, low-affinity variants of splitFAST, and studied the dynamics of different MCSs at high spatiotemporal resolution, both in vitro and in vivo. We demonstrated that these versatile reporters suit different experimental setups well, allowing one to address challenging biological questions. Using these novel probes, we identified a hitherto unknown pathway, in which calcium (Ca2+) signalling dynamically regulates endoplasmic reticulum-mitochondria juxtaposition, characterizing the underlying mechanism. Finally, by integrating Ca2+-sensing capabilities into the splitFAST technology, we introduced PRINCESS (PRobe for INterorganelle Ca2+-Exchange Sites based on SplitFAST), an unprecedented class of reporters to simultaneously detect MCSs and measure the associated Ca2+ dynamics using a single biosensor.

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Garcia Casas, P., Rossini, M., Pavenius, L., Saeed, M., Arnst, N., Sonda, S., Bruzzone, M., Berno, V., Raimondi, A., Sassano, M. L., Naia, L., Agostinis, P., Sturlese, M., Niemeyer, B. A., Brismar, H., Ankarcrona, M., Gautier, A., Pizzo, P., Filadi, R.. 2023-12-28. Simultaneous detection of membrane contact dynamics and associated Ca2+ signals by reversible chemogenetic reporters. https://doi.org/10.1101/2023.12.28.573515

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