HTD2 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from human embryonic kidney HEK293T cells. This product enables loss-of-function studies of the HTD2 gene, which encodes a mitochondrial enzyme. The polyclonal population was generated via CRISPR/Cas9-mediated gene disruption, resulting in a heterogeneous pool of edited cells that lack functional HTD2 expression. This format provides a robust model for investigating HTD2-dependent processes without clonal selection biases.
The host cell line, HEK293T, is a derivative of HEK293 cells that stably expresses the SV40 large T antigen, enhancing episomal plasmid replication and conferring high transfection efficiency. Originally from human embryonic kidney, HEK293T cells are epithelial and widely used for transient expression, lentivirus production, and functional assays. Their rapid growth and adaptability make them ideal for generating gene-edited models to explore mitochondrial and metabolic pathways.
HTD2 encodes a mitochondrial 3-hydroxyacyl-thioester dehydratase that operates within the mitochondrial fatty acid synthesis (mtFAS) pathway. It catalyzes the dehydration of 3-hydroxyacyl-ACP to enoyl-ACP, a key step preceding reduction by mitochondrial enoyl-CoA reductase (MECR). This reaction is essential for producing octanoyl-ACP, the direct precursor for lipoic acid biosynthesis. HTD2 functions in concert with MCAT, OXSM, and MECR. Upstream, HTD2 expression is regulated by PGC-1??, NRF1, TFAM, and PPAR??, which coordinate mitochondrial biogenesis and metabolic gene expression. Downstream, its activity impacts lipoic acid levels and iron-sulfur cluster assembly, connecting mtFAS to mitochondrial dehydrogenase function.
Disruption of HTD2 in HEK293T cells provides a physiologically relevant model for studying mitochondrial dysfunction associated with neurometabolic disorders. HTD2 mutations have been linked to mitochondrial disease, epilepsy, and developmental delay, underscoring its importance in neural metabolism. Although HEK293T cells are kidney-derived, they retain a functional mitochondrial network and are suitable for dissecting fundamental mitochondrial processes. This knockout model allows researchers to examine the consequences of impaired mtFAS on cellular metabolism, lipoic acid availability, and downstream pathways in a tractable in vitro system.
Researchers can employ HTD2 Knockout HEK293T Polyclonal Cells in diverse experimental applications, including profiling mitochondrial fatty acids by mass spectrometry, quantifying lipoic acid levels, and measuring oxygen consumption rates to assess respiratory chain activity. The cells are valuable for investigating HTD2-dependent signaling, testing chemical modulators in drug screening assays, and performing metabolic flux analyses. Additional techniques such as Western blotting for HTD2, RT-qPCR analysis of mitochondrial genes, and immunofluorescence imaging of mitochondrial morphology can be utilized. This knockout model is an essential tool for advancing understanding of mtFAS and lipoic acid biology. For further information, please contact Ascent Research.
