The ECI2 Knockout HEK293T Polyclonal Cells product provides a ready-to-use polyclonal population of HEK293T cells that have undergone CRISPR/Cas9-mediated disruption of the ECI2 gene. This gene-edited pool serves as a loss-of-function model for studying the role of enoyl-CoA delta isomerase 2 in cellular metabolism. As a polyclonal knockout population, it offers a heterogeneous genetic background while maintaining null expression of the target gene, suitable for bulk assays and functional studies.
The parental HEK293T cell line is a widely utilized human embryonic kidney epithelial cell line, originally derived by transformation with adenovirus 5 DNA and stably expressing the SV40 large T antigen. This cell line is highly transfectable and supports robust protein expression and viral production, making it an ideal host for gene-editing applications. Its epithelial origin and immortalized nature facilitate consistent cell culture and reproducible experimental outcomes, while its well-characterized metabolism provides a suitable backdrop for investigating mitochondrial fatty acid oxidation pathways.
ECI2 encodes a mitochondrial enzyme essential for the isomerization of 3-cis and 3-trans enoyl-CoA esters to their 2-trans forms, a critical step in the beta-oxidation of unsaturated fatty acids. Functioning downstream of PPARalpha and nutritional cues monitored by the insulin/glucagon axis, ECI2 acts in concert with long-chain acyl-CoA dehydrogenase (ACADVL) and the mitochondrial trifunctional protein subunits alpha (HADHA) and beta (HADHB), as well as short-chain enoyl-CoA hydratase (ECHS1), to facilitate complete fatty acid degradation. The enzyme??s isomerization activity yields 2-trans enoyl-CoA intermediates that enter the beta-oxidation spiral, ultimately generating acetyl-CoA units for ATP synthesis via oxidative phosphorylation. Disruption of ECI2 impairs this sequential process, leading to a block in unsaturated fatty acid catabolism and reduced energy production from lipid substrates.
Ablation of ECI2 in HEK293T cells creates a metabolic model that recapitulates key features of genetic defects in mitochondrial unsaturated fatty acid oxidation. In these polyclonal knockout cells, the absence of functional enoyl-CoA delta isomerase 2 activity results in the accumulation of 3-cis/3-trans enoyl-CoA species, a hallmark of certain fatty acid oxidation disorders. This metabolic bottleneck also diminishes acetyl-CoA and ATP production, rendering the cells more reliant on glycolysis and other non-lipid energy sources. Consequently, the knockout model provides a physiologically relevant system for investigating the metabolic adaptations and compensatory pathways that arise when mitochondrial lipid catabolism is compromised.
Researchers can employ ECI2 Knockout HEK293T Polyclonal Cells to dissect the role of unsaturated fatty acid oxidation in cell physiology, including its impact on energy homeostasis, mitochondrial respiration, and lipid intermediate signaling. The cells are compatible with a range of experimental approaches, such as quantitative PCR and western blotting to verify ECI2 disruption and downstream protein expression changes, Seahorse metabolic flux analysis to assess oxidative phosphorylation and glycolytic parameters, and targeted metabolomics to profile acyl-CoA esters and other lipid intermediates. Additional applications include evaluating pharmacological rescue agents or gene therapy vectors for metabolic disorders, and investigating the interplay between fatty acid oxidation and other metabolic pathways. For further information, please contact Ascent Research.
