@article {Chang-Graham627877, author = {Alexandra L. Chang-Graham and Jacob L. Perry and Alicia C. Strtak and Nina K. Ramachandran and Jeanette M. Criglar and Asha A. Philip and John T. Patton and Mary K. Estes and Joseph M. Hyser}, title = {Rotavirus Calcium Dysregulation Manifests as Dynamic Calcium Signaling in the Cytoplasm and Endoplasmic Reticulum}, elocation-id = {627877}, year = {2019}, doi = {10.1101/627877}, publisher = {Cold Spring Harbor Laboratory}, abstract = {Like many viruses, rotavirus (RV) dysregulates calcium homeostasis by elevating cytosolic calcium ([Ca2+]cyt) and decreasing endoplasmic reticulum (ER) stores. While an overall, monophasic increase in [Ca2+]cyt during RV infection has been shown, the nature of the RV-induced aberrant calcium signals and how they manifest over time at the single-cell level have not been characterized. Thus, we generated cell lines and human intestinal enteroids (HIEs) stably expressing cytosolic and/or ER-targeted genetically-encoded calcium indicators to characterize calcium signaling throughout RV infection by time-lapse imaging. We found that RV induces highly dynamic [Ca2+]cyt signaling that manifest as hundreds of discrete [Ca2+]cyt spikes, which increase during peak infection. Knockdown of nonstructural protein 4 (NSP4) attenuates the [Ca2+]cyt spikes, consistent with its role in dysregulating calcium homeostasis. RV-induced [Ca2+]cyt spikes were primarily from ER calcium release and were attenuated by inhibiting the store-operated calcium entry (SOCE) channel Orai1. RV-infected HIEs also exhibited prominent [Ca2+]cyt spikes that were attenuated by inhibiting SOCE, underlining the relevance of these [Ca2+]cyt spikes to gastrointestinal physiology and role of SOCE in RV pathophysiology. Thus, our discovery that RV increases [Ca2+]cyt by dynamic Ca2+ signaling, establishes a new, paradigm-shifting understanding of the spatial and temporal complexity of virus-induced Ca2+ signaling.}, URL = {https://www.biorxiv.org/content/early/2019/05/04/627877}, eprint = {https://www.biorxiv.org/content/early/2019/05/04/627877.full.pdf}, journal = {bioRxiv} }