The ACACA Knockout HCT 116 Polyclonal Cells product consists of a CRISPR/Cas9-edited polyclonal population of HCT 116 cells with targeted disruption of the ACACA locus. As a polyclonal pool, this model comprises a heterogeneous mix of knockout cells, enabling functional studies of acetyl-CoA carboxylase alpha (ACC1) without single-cell clonal expansion. Disruption of ACACA abrogates de novo fatty acid synthesis by eliminating the conversion of acetyl-CoA to malonyl-CoA, thereby furnishing a loss-of-function tool for metabolic investigations.
The parental HCT 116 line is a human colorectal carcinoma model featuring a KRAS G13D mutation, microsatellite instability (MSI), and loss of MLH1 expression, along with mutations in TP53 and CTNNB1. These genetic alterations render HCT 116 cells particularly valuable for studying metabolic dependencies driven by oncogenic KRAS and mismatch repair deficiency, establishing a clinically relevant platform for dissecting lipid anabolism in colon cancer.
ACACA encodes ACC1, the rate-limiting enzyme of de novo fatty acid synthesis, which carboxylates acetyl-CoA to malonyl-CoA in a biotin-dependent reaction. Malonyl-CoA serves as the substrate for fatty acid synthase (FASN) and allosterically inhibits carnitine palmitoyltransferase 1 (CPT1), thereby suppressing fatty acid oxidation. ACC1 is phosphorylated and inhibited by AMPK at Ser79, while insulin and SREBP1 upregulate its expression. Protein phosphatase 2A (PP2A) reverses AMPK-mediated inhibition. Interacting cofactors include acetyl-CoA, citrate, and ATP-citrate lyase. Knockout of ACACA therefore disrupts both lipogenesis and the malonyl-CoA?CCPT1 regulatory node, profoundly altering cellular lipid metabolism and energy homeostasis.
In HCT 116 cells, ACACA knockout eliminates endogenous fatty acid synthesis, forcing reliance on exogenous lipids and potentially unveiling synthetic lethal vulnerabilities with the KRAS G13D oncogene, which is known to drive lipogenic reprogramming. This model is ideally suited for investigating the metabolic adaptations of colorectal cancers with MSI and KRAS activation, and for assessing the therapeutic potential of ACC1 inhibitors in a defined genetic background.
Applications include metabolic flux analysis via [14C]-acetate incorporation, lipidomic profiling by mass spectrometry, and Seahorse assays for fatty acid oxidation. The polyclonal pool supports western blot profiling of ACC1 and phospho-ACC1 (Ser79) to evaluate AMPK regulation, as well as RT-qPCR and ChIP-qPCR to probe SREBP1-mediated transcription. It is also valuable for inhibitor screening, synthetic lethal interaction studies with KRAS-targeted agents, and RNA-seq transcriptomics. For further details or custom applications, please contact Ascent Research.
