@article {70083,
	title = {An expanded palette of bright and photostable organellar Ca2+ sensors},
	journal = {eLife},
	year = {2025},
	month = {2025 Aug 26},
	abstract = {<p>The use of fluorescent sensors for functional imaging has revolutionized the study of organellar Ca2+\&nbsp;signaling. However, understanding the dynamic interplay between intracellular Ca2+\&nbsp;sinks and sources has been hindered by the lack of bright, photostable, and multiplexed measurements in different organelles, limiting our ability to define how Ca2+\&nbsp;shapes cell physiology across fields of biology. Here we introduce a new toolkit of chemigenetic organellar Ca2+\&nbsp;indicators whose color is tunable by reconstituting their fluorescence with different exogenous rhodamine dye-ligands, which significantly expand the capacity for multiplexing organellar Ca2+\&nbsp;measurements. These sensors, which we named ER-HaloCaMP and Mito-HaloCaMP, are optimized to report Ca2+dynamics in the endoplasmic reticulum (ER) and mitochondria of mammalian cells and neurons, and show significantly improved brightness, photostability and responsiveness when compared to current best-in-class alternatives. Using either red or far-red dye-ligands, both ER-HaloCaMP and Mito-HaloCaMP enable visualizing ER and mitochondrial Ca2+\&nbsp;dynamics in neuronal axons, a subcellular location that only contains a few ER tubules and small mitochondria, structural limitations that have impaired measurements with previous red sensors. To show the expanded multiplexing capacities of our toolkit, we measured interorganellar Ca2+\&nbsp;fluxes simultaneously in three different subcellular compartments in live cells, revealing that the amplitude of ER Ca2+release controls the efficacy of ER-mitochondria Ca2+\&nbsp;coupling in a cooperative manner. Organellar HaloCaMPs enable also measuring Ca2+\&nbsp;dynamics in intact brain tissue from flies and rodents, demonstrating their versatility across biological models. Our new toolkit provides an expanded palette of bright, photostable and responsive organellar Ca2+\&nbsp;sensors, which will facilitate future studies of intracellular Ca2+\&nbsp;signaling across fields of biology in health and disease.</p>
},
	doi = {10.7554/elife.107845.1},
	url = {http://dx.doi.org/10.7554/eLife.107845.1},
	author = {Moret, Agathe and Farrants, Helen and Fan, Ruolin and Zingg, Kelsey G and Silva, Bryon and Roselli, Camilla and Oertner, Thomas G and Gee, Christine E and Hadjieconomou, Dafni and Rangaraju, Vidhya and Schreiter, Eric R and de Juan-Sanz, Jaime}
}
