The ACOT7 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout population engineered to disrupt the ACOT7 gene in human HEK293T cells. This pooled model provides a system to study mitochondrial acyl-CoA thioesterase function without clonal isolation, preserving population heterogeneity while ablating target-gene expression via CRISPR/Cas9-mediated disruption.
HEK293T cells, a derivative of HEK293 transformed with adenovirus 5 DNA, stably express the SV40 large T antigen, enabling episomal replication of plasmids with the SV40 origin. As human embryonic kidney epithelial cells, they are widely used for protein expression, viral production, and gene-editing applications due to high transfectability and robust metabolic activity.
ACOT7 encodes a mitochondrial acyl-CoA thioesterase that hydrolyzes long-chain acyl-CoAs (e.g., palmitoyl-CoA) to free fatty acids and CoA, thereby regulating metabolite pools. Its expression is controlled by PPAR??, SREBP-1c, and PGC-1??, integrating fatty acid metabolism, CoA biosynthesis, and energy homeostasis. ACOT7 acts downstream of long-chain acyl-CoA synthetases, upstream of CPT1, and interacts with ??-oxidation enzymes, electron transport chain complexes, and CoA-binding proteins. Knockout is expected to lead to acyl-CoA accumulation, reduced free fatty acid and CoA levels, impaired ??-oxidation, and altered lipid signaling, impacting cellular energetics and mitochondrial function.
In HEK293T cells, ACOT7 loss disrupts mitochondrial lipid handling, offering a model to probe lipid metabolism?Cenergy production crosstalk. Functional assays such as Seahorse metabolic flux analysis, fatty acid oxidation measurements, and mass spectrometry lipidomics quantify changes in respiration, lipid utilization, and acyl-CoA/free fatty acid profiles. These approaches connect ACOT7 activity to phenotypes in metabolic disorders like obesity, NAFLD, and cancer, where thioesterase regulation is increasingly recognized.
Applications span fatty acid metabolism studies, CoA homeostasis research, mitochondrial function analysis, and metabolic disease modeling. Gene disruption can be verified by RT-qPCR and Western blotting, while mitochondrial morphology is assessed via immunofluorescence. The model supports drug screening for lipid metabolism disorders and investigation of PPAR??/PGC-1?? transcriptional responses to altered lipid and CoA status. For more information, contact Ascent Research.
