The HRAS Knockout A-549 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population engineered to disrupt the HRAS gene in the A-549 human lung adenocarcinoma cell line. This heterogeneous pool provides a loss-of-function model that avoids clonal selection biases, suitable for bulk population studies. Frozen stocks of early-passage cells are supplied for direct culture and experimental use.
The A-549 cell line originated from a 58-year-old Caucasian male lung carcinoma and displays adherent epithelial morphology typical of alveolar basal epithelial cells. It harbors a KRAS G12S mutation, serving as a key model for lung adenocarcinoma. This background enables analysis of HRAS-specific signaling in the presence of oncogenic KRAS, facilitating studies of isoform dependency and compensation.
HRAS encodes a small GTPase functioning as a molecular switch, cycling between inactive GDP-bound and active GTP-bound conformations. Upstream, receptor tyrosine kinases such as EGFR and FGFR respond to growth factors (e.g., EGF, FGF) by recruiting GRB2-SOS1 complexes, which catalyze nucleotide exchange on HRAS. Active, GTP-loaded HRAS binds and activates multiple downstream effectors, notably RAF kinases (BRAF, CRAF), which then phosphorylate MEK1/2 and ERK1/2, culminating in phosphorylation of transcription factors like ELK1 and promoting cell cycle progression. In parallel, HRAS activates PI3K, leading to AKT and mTOR signaling that enhances cell survival and metabolism. Signaling is terminated by GTPase-activating proteins (GAPs) including NF1 and p120GAP, which accelerate GTP hydrolysis. Thus, HRAS integrates proliferative and survival signals downstream of extracellular stimuli.
In A-549 cells, HRAS knockout allows dissection of RAS signaling contributions in lung adenocarcinoma. Despite predominant KRAS G12S signaling, HRAS may drive parallel or compensatory pathways, and its disruption is anticipated to reduce MAPK and PI3K pathway activity, impairing proliferation, survival, and anchorage-independent growth. This model supports research into RAS isoform addiction, resistance mechanisms to KRAS inhibitors, and the role of HRAS in RASopathies.
Typical applications include Western blotting and RT-qPCR for confirming HRAS loss and assessing downstream targets; cell proliferation (MTT, BrdU) and colony formation assays; soft agar assays for anchorage-independent growth; xenograft tumor studies; flow cytometry for cell cycle and apoptosis; and phospho-signaling analysis. The product is useful for drug sensitivity testing and validation of RAS pathway inhibitors. For further details or custom cell engineering, please contact Ascent Research.
