The ATP2A1 Knockout HEK293T Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout cell population derived from HEK293T cells, in which the ATP2A1 gene encoding sarco/endoplasmic reticulum Ca2+-ATPase 1 (SERCA1) has been disrupted. This loss-of-function model provides a pooled population of edited cells, enabling robust analysis of SERCA1-dependent calcium handling without monoclonal selection, and is ideal for studying the impact of ATP2A1 ablation on intracellular calcium dynamics and downstream signaling networks.
HEK293T cells are a widely used human embryonic kidney epithelial cell line that stably expresses the SV40 large T antigen. This property enhances episomal replication of plasmids containing the SV40 origin of replication, making HEK293T cells highly efficient for transient protein expression, lentiviral packaging, and gene editing applications. Their robust growth and ease of transfection have established HEK293T as a versatile host for functional genomics studies, including CRISPR-based knockout screening, where the polyclonal format maintains population-level diversity while simplifying experimental workflows.
ATP2A1 encodes SERCA1, the primary ATPase responsible for transporting cytosolic Ca2+ into the endoplasmic and sarcoplasmic reticulum lumen, a process essential for muscle relaxation and cellular calcium homeostasis. SERCA1 activity is regulated by interacting factors phospholamban (PLN) and sarcolipin (SLN), and its expression is controlled by upstream transcription factors such as MyoD and MEF2C, with further modulation by CaMKII and calcineurin. Downstream, SERCA1-dependent calcium stores influence the activation of NFATC1 and CAMK2A, linking Ca2+ flux to transcriptional responses. Disruption of ATP2A1 in this model perturbs these interconnected pathways, potentially triggering ER stress and altering autophagy-related genes, thereby providing a platform to dissect calcium signaling networks.
In the HEK293T cellular context, ATP2A1 knockout interrupts the normal reuptake of cytosolic Ca2+ into the ER, leading to altered calcium storage and dysregulated calcium-dependent signaling. Although HEK293T cells are not of muscle origin, they express key components of calcium handling machinery, and SERCA1 loss can unmask compensatory mechanisms or reveal non-muscle roles of SERCA isoforms. This knockout model therefore allows investigation of ER calcium depletion, unfolded protein response activation, and the interplay between Ca2+ signaling and cell survival pathways in an easily manipulable epithelial cell background, aiding in the dissection of fundamental calcium biology without muscle-specific confounding factors.
Researchers can employ this polyclonal knockout population in a variety of experimental contexts, including calcium imaging using Fluo-4 to measure cytosolic and ER calcium dynamics, RT-qPCR and western blotting to assess expression changes in SERCA interactors and downstream effectors, and flow cytometry to quantify calcium flux under agonist stimulation. The model is particularly suited for Brody disease research, muscle physiology studies even in a non-muscle host, and high-throughput screening of SERCA modulators. By combining cell viability assays under calcium stress with targeted pathway analysis, users can explore ATP2A1-dependent mechanisms in health and disease. For further details or technical inquiries, please contact Ascent Research.
