The Golgi calcium ATPase pump plays an essential role in AAV trafficking and transduction.

Adeno-associated viruses (AAVs) are dependoparvoviruses that have proven useful for therapeutic gene transfer; however, our understanding of host factors that influence AAV trafficking and transduction is still evolving. Here, we investigate the role of cellular calcium in the AAV infectious pathway. First, we demonstrate a critical role for the host Golgi-resident ATP-powered calcium pump (secretory pathway calcium ATPase 1; SPCA1) encoded by the ATP2C1 gene in AAV infection. CRISPR-based knockout (KO) of ATP2C1 decreases transduction by different AAV serotypes. ATP2C1 KO does not appear to inhibit AAV binding, cellular uptake or nuclear entry; however, capsids within ATP2C1 KO cells demonstrate disperse and punctate trafficking distinct from the perinuclear, trans -Golgi pattern observed in normal cells. In addition, we observed a defect in the ability of AAV capsids to undergo conformational changes and support efficient vector genome transcription in ATP2C1 KO cells. The calcium chelator, BAPTA-AM, which reduces cytosolic calcium, rescues the defective ATP2C1 KO phenotype and AAV transduction in vitro Conversely, the calcium ionophore, Ionomycin, which disrupts calcium gradients, blocks AAV transduction. Further, we demonstrate that modulating calcium in the murine brain using BAPTA-AM augments AAV gene expression in vivo. Taken together, we postulate that the maintenance of an intracellular calcium gradient by the calcium ATPase and processing within the Golgi compartment are essential for priming the capsid to support efficient AAV genome transcription. IMPORTANCE Adeno-associated viruses (AAVs) have proven to be effective gene transfer vectors. However, our understanding of how the host cell environment influences AAV transduction is still evolving. In the present study, we investigate the role of ATP2C1 , which encodes a membrane calcium transport pump, SPCA1, essential for maintaining cellular calcium homeostasis on AAV transduction. Our results indicate that cellular calcium is essential for efficient intracellular trafficking and conformational changes in the AAV capsid that support efficient genome transcription. Further, we show that pharmacological modulation of cellular calcium levels can potentially be applied to improve the AAV gene transfer efficiency.