The ITPKC Knockout A-549 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the A-549 human lung adenocarcinoma epithelial cell line. This polyclonal pool carries targeted disruptions in the ITPKC gene locus, introduced via CRISPR/Cas9-mediated genome editing, resulting in a loss-of-function model for inositol-trisphosphate 3-kinase C. The heterogeneous population enables study of ITPKC function without clonal selection artifacts, preserving genetic diversity typical of cancer cell research.
The parental A-549 cell line was originally isolated from a 58-year-old Caucasian male with lung carcinoma and serves as a widely used model of lung adenocarcinoma. These cells exhibit characteristics of alveolar type II pneumocytes and are routinely applied in respiratory epithelial biology, oncology, and drug discovery. Their adherent growth, stable karyotype, and well-documented signaling networks make them a robust platform for genetic manipulation and functional genomics studies.
ITPKC encodes inositol-trisphosphate 3-kinase C, a kinase that phosphorylates the second messenger inositol 1,4,5-trisphosphate (IP3) to inositol 1,3,4,5-tetrakisphosphate (IP4), thereby dampening IP3-dependent calcium release from intracellular stores. In the context of T cell receptor (TCR) signaling, ITPKC acts downstream of the TCR/CD3 complex, ZAP70, LAT, and PLCG1, and functions as a negative regulator of calcium signaling. By reducing cytoplasmic calcium levels, ITPKC limits calmodulin-mediated activation of calcineurin, which in turn decreases dephosphorylation and nuclear translocation of NFAT transcription factors. This cascade ultimately attenuates the transcriptional induction of cytokines such as IL-2.
Although ITPKC has been extensively studied in lymphocytes, its expression and function in lung epithelial cells are less characterized. The ITPKC knockout A-549 model provides a unique tool to investigate how inositol phosphate metabolism influences calcium-dependent processes in lung adenocarcinoma cells. Given that A-549 cells retain certain signaling modules common to immune cells, including calcium flux and NFAT-dependent gene expression, this model enables dissection of ITPKC??s role in epithelial inflammation, cytokine production, and potentially tumor microenvironment interactions. Disruption of ITPKC in this background could reveal novel links between phosphoinositide signaling and lung cancer progression, such as altered proliferation, migration, or inflammatory mediator secretion.
Researchers can employ this polyclonal knockout pool in a variety of functional assays. Loss of ITPKC protein can be confirmed via Western blotting, and genomic editing verified by Sanger sequencing. The impact on calcium dynamics is assessable through intracellular calcium flux assays using Fluo-4 AM dye. Transcriptional consequences can be measured by NFAT luciferase reporter assays or RT-qPCR profiling of downstream cytokine genes. This model is suitable for screening small-molecule kinase inhibitors that may modulate IP3 metabolism, and for phenotypic analyses including cell proliferation (MTT), migration, and invasion. For further information, please contact Ascent Research.
