The ATP5F1A Knockout HAP1 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout cell population, generated by disrupting the ATP5F1A gene in the near-haploid HAP1 cell line. This product offers a heterogeneous pool of edited cells, each carrying loss-of-function mutations in the nuclear gene encoding the alpha subunit of mitochondrial ATP synthase. The polyclonal format ensures reproducibility and is ideal for large-scale screening applications, avoiding the artifacts of single-clone expansion.
HAP1 cells are a near-haploid human cell line derived from KBM-7 chronic myeloid leukemia cells, exhibiting a fibroblast-like morphology and male karyotype. Their near-haploid genome simplifies CRISPR/Cas9-mediated gene disruption, as alteration of a single allele yields a functional knockout. Widely employed in genetic screens and functional genomics, HAP1 cells provide an ideal system for studying gene function, particularly in pathways where heterozygous effects would confound interpretation.
ATP5F1A encodes the alpha subunit of the F1 catalytic domain of mitochondrial ATP synthase (Complex V), essential for ATP production via oxidative phosphorylation. Its expression is regulated by PGC-1??, NRF1, and TFAM, linking mitochondrial biogenesis to ATP synthase assembly. The alpha subunit interacts with ATP5F1B, ATP5O, and the inhibitory factor IF1 to form the F1 complex, which couples the proton gradient generated by electron transport chain complexes I?CIV and cytochrome c to ATP synthesis. ATP5F1A disruption abolishes ATP synthase activity, collapses mitochondrial membrane potential, reduces cellular ATP levels, and increases reactive oxygen species production, thereby modeling mitochondrial energy failure and oxidative stress.
In HAP1 cells, ATP5F1A knockout induces profound mitochondrial dysfunction mimicking diseases such as Leigh syndrome and various neurodegenerative disorders. The near-haploid background ensures a rapid and complete loss of ATP synthase activity, leading to reliance on glycolysis and metabolic reprogramming. This model is well-suited for assessing mitochondrial membrane potential changes with JC-1 or TMRM, measuring oxygen consumption via Seahorse analysis, and studying the interplay between energy metabolism and mitochondrial quality control pathways.
Typical applications include mitochondrial dysfunction modeling, drug screening for ATP synthase modulators, and investigation of energy metabolism. Researchers can validate gene disruption by Western blotting for ATP5F1A, quantify ATP levels using luciferase assays, and profile mitochondrial gene expression by RT-qPCR. The polyclonal population is ideal for cell viability assays and high-throughput drug screens. For additional customization or monoclonal isolation, please contact Ascent Research.
