TMEM63A Knockout COS1 Cell Line

The TMEM63A Knockout COS1 Cell Line is a CRISPR/Cas9-mediated knockout model in the COS1 cell line, designed to disrupt the TMEM63A gene encoding a mechanosensitive calcium-permeable cation channel. This channel is critical for cell volume regulation and osmotic stress responses. The stable knockout cell line provides a defined loss-of-function system for investigating mechanotransduction and calcium signaling pathways without the variability of transient gene silencing.

COS1 cells originate from CV-1 African green monkey kidney fibroblasts (Chlorocebus aethiops) and are immortalized with SV40 large T antigen. They are widely employed for transient protein expression due to their ability to replicate plasmids bearing the SV40 origin. As a kidney fibroblast model, COS1 cells retain relevant ion channel and volume regulatory mechanisms, making them suitable for studying mechanosensitive channels and osmotic stress signaling, with robust and rapid proliferation.

TMEM63A is activated by mechanical stretch, osmotic stress, and cell swelling, mediating calcium influx that triggers downstream signaling cascades. This calcium entry activates calmodulin and calcineurin, which dephosphorylate NFAT transcription factors, promoting their nuclear translocation. Concurrently, TMEM63A-mediated calcium signals modulate the Hippo pathway, affecting YAP and TAZ activity. The channel physically and functionally interacts with the actin cytoskeleton and calcium-calmodulin-dependent kinases, integrating mechanical cues with transcriptional responses. Disruption of TMEM63A abrogates these mechanotransduction pathways, providing a clean background for mechanistic dissection.

In COS1 kidney fibroblasts, TMEM63A knockout impairs normal mechanosensitive calcium entry, likely disrupting regulatory volume decrease and hypotonic stress responses. This model is valuable for investigating TMEM63A-related pathologies, including cerebellar ataxia, cancer, and neurological disorders. The renal fibroblast background enables focused study of volume regulation and mechanotransduction, while the cell line’s robust growth supports high-throughput drug screening and functional genomics approaches targeting channelopathies.

The cell line supports calcium imaging and patch-clamp electrophysiology to assess ion channel activity, cell volume assays for osmoregulation, and immunofluorescence for actin stress fiber organization. Western blotting detects YAP phosphorylation, and qRT-PCR quantifies downstream gene expression changes; cell migration assays evaluate functional outcomes. These applications facilitate drug target validation for mechanosensitive channels and the development of disease models in neuroscience and oncology. For further details or technical assistance, please contact Ascent Research.