The ATP2B4 Knockout HT29 Polyclonal Cells represent a CRISPR/Cas9-edited HT29 cell population designed to disrupt the ATP2B4 gene, thereby eliminating functional expression of the plasma membrane Ca2+ ATPase PMCA4. This polyclonal knockout pool comprises a heterogeneous mixture of cells with distinct editing events, collectively ensuring robust loss of PMCA4 activity while avoiding clonal selection biases. The model provides a physiologically relevant context to examine PMCA4-dependent processes in colorectal cancer.
HT29 cells originate from a primary colorectal adenocarcinoma and maintain differentiated epithelial characteristics, including the expression of villin and mucins, as well as the capacity for enterocytic differentiation. They are widely employed as an intestinal epithelial model in colorectal cancer research, suitable for investigating tumorigenic signaling, drug metabolism, and metastasis.
PMCA4, a P-type IIB ATPase encoded by ATP2B4, is a high-affinity, low-capacity calcium pump critical for fine-tuning cytosolic calcium levels. Its activity is modulated by Ca2+/calmodulin, PKA, PKC, and caspase-mediated cleavage. PMCA4 forms complexes with scaffolding proteins such as NHERF2, ??1-syntrophin, nNOS, and Homer, thereby localizing calcium extrusion to specific membrane microdomains. Through these interactions, PMCA4 regulates downstream effectors including calcineurin, NFAT transcription factors, and TRP ion channels, exerting control over pathways like Wnt, PI3K/AKT, and MAPK/ERK. This positions PMCA4 at the nexus of calcium-dependent signal transduction, influencing gene expression, proliferation, and apoptotic decisions.
In HT29 colorectal adenocarcinoma cells, PMCA4-mediated calcium homeostasis is integral to balancing cell cycle progression and apoptosis. Disruption of ATP2B4 in this polyclonal model elevates intracellular calcium, thereby altering calcineurin-NFAT and PI3K/AKT pathway activities. The resulting dysregulation can suppress cell growth and promote cell death, underscoring PMCA4??s potential tumor-suppressive functions. This model is valuable for dissecting calcium signaling contributions to colorectal cancer pathology.
Researchers can employ this polyclonal knockout for detailed functional analyses, including live-cell calcium imaging with Fluo-4 or Fura-2 indicators, flow cytometric assessment of apoptosis, MTT-based viability assays, and Transwell migration/invasion experiments. The model supports molecular characterization via RT-qPCR, Western blotting, RNA-seq, and immunofluorescence to validate pathway alterations. It is also ideal for screening pharmacological modulators of calcium transport and for studying cross-talk between calcium signaling and oncogenic cascades. For technical support or to request a quote, please contact Ascent Research.
