The GRAMD1C Knockout NCI-H1975 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal cell population derived from the NCI-H1975 lung adenocarcinoma cell line, engineered to disrupt the GRAMD1C gene. This knockout model provides a comprehensive loss-of-function tool for investigating GRAMD1C-dependent cholesterol transport and signaling in a human cancer context, without relying on clonal selection, thus preserving population-level heterogeneity.
NCI-H1975 cells are an established model of human lung adenocarcinoma derived from a female donor, harboring an activating EGFR L858R mutation and retaining sensitivity to tyrosine kinase inhibitors (TKIs). These cells are widely employed to study EGFR-driven oncogenic signaling pathways, including downstream AKT/mTOR and RAS/MAPK cascades, and to evaluate drug responses in non-small cell lung cancer (NSCLC).
GRAMD1C, also known as Aster-C, is a cholesterol transporter that localizes to endoplasmic reticulum?Cplasma membrane (ER?CPM) contact sites, where it facilitates non-vesicular cholesterol transfer from the PM to the ER. Its activity is regulated by intracellular cholesterol levels via the SREBP2 and LXR transcriptional programs. GRAMD1C interacts with VAPA/VAPB at the ER and binds phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) at the PM, coordinating with OSBP in sterol sensing and transport. Loss of GRAMD1C disrupts cholesterol distribution, leading to altered processing of SREBP transcription factors, impaired AKT membrane recruitment, and dysregulated mTORC1 signaling, collectively impacting lipid homeostasis and cell growth control.
In the NCI-H1975 lung adenocarcinoma model, GRAMD1C knockout enables dissection of cholesterol-mediated regulation of oncogenic signaling. The EGFR L858R mutation drives downstream pathways that are sensitive to membrane cholesterol levels, including AKT/mTOR, which relies on cholesterol-rich lipid rafts for proper signal transduction. By depleting GRAMD1C, researchers can examine how ER?CPM cholesterol flux influences EGFR signaling, SREBP-dependent lipogenesis, and tumor cell proliferation, providing insights into lipid metabolic reprogramming in NSCLC. This knockout model is particularly valuable for evaluating GRAMD1C as a node connecting cholesterol homeostasis to cancer cell growth and survival.
Researchers can employ this polyclonal knockout population to examine cholesterol trafficking via BODIPY-cholesterol assays, quantify cellular cholesterol, and assess signaling through phospho-pAKT and p-S6 analysis. Proximity ligation assays validate ER?CPM contact site disruption, while cell proliferation assays gauge growth effects and RNA-seq reveals transcriptomic changes. These applications support investigating GRAMD1C in EGFR-driven tumorigenesis and its potential as a therapeutic target in lipid metabolic reprogramming. For further details, please contact Ascent Research.
