The L2HGDH Knockout HAP1 Polyclonal Cells product is a CRISPR/Cas9-edited polyclonal knockout cell population designed to disrupt the human L2HGDH gene in the near-haploid HAP1 cell line. This loss-of-function model, generated by CRISPR/Cas9-mediated gene disruption, comprises a heterogeneous pool of cells carrying L2HGDH knockout alleles, enabling robust functional studies without clonal isolation. The polyclonal format mitigates clonal variation and provides a representative knockout background for interrogating L-2-hydroxyglutarate metabolism and downstream signaling pathways.
HAP1 is a near-haploid human cell line derived from the chronic myeloid leukemia cell line KBM-7, established from a male patient. Its near-haploid karyotype simplifies genetic analyses, making it a well-established host for functional genomics, genetic screening, and haploid genetics. The limited gene copy number facilitates straightforward genotype?Cphenotype correlations and enhances the efficiency of CRISPR-based gene disruption, rendering HAP1 an ideal background for studying genes involved in metabolic and epigenetic regulation.
L2HGDH encodes a mitochondrial dehydrogenase that catalyzes the oxidation of the oncometabolite L-2-hydroxyglutarate (L-2HG) to alpha-ketoglutarate (??-KG), thereby preventing competitive inhibition of ??-KG-dependent dioxygenases. In this knockout model, loss of L2HGDH leads to intracellular L-2HG accumulation, which in turn inhibits TET family DNA demethylases and JmjC domain-containing histone demethylases, among other dioxygenases. Upstream, L-2HG is generated by lactate dehydrogenase A (LDHA) under normoxic and hypoxic conditions. Disruption of L2HGDH thus dysregulates DNA and histone methylation patterns by impairing ??-KG-dependent enzymatic activities, with particular impact on TET2-mediated DNA hydroxymethylation and JmjC-mediated histone demethylation.
In the HAP1 cellular context, near-haploidy offers distinct advantages for dissecting L2HGDH function. Because the cell line carries only one copy of most chromosomes, the knockout phenotype is directly attributable to L2HGDH disruption without interference from a second wild-type allele. This simplifies the interpretation of L-2HG accumulation, epigenetic changes, and metabolic alterations. The HAP1 background is particularly suited for investigating cancer metabolism and neurometabolic disorders, as L2HGDH mutations are linked to L-2-hydroxyglutaric aciduria and tumorigenesis. The polyclonal knockout population enables high-throughput screening and bulk profiling while minimizing the confounding effects of individual clonal selection.
Researchers can employ this knockout model in a wide array of experimental applications, including quantitative measurement of L-2HG by LC-MS, analysis of DNA hydroxymethylation and histone methylation via ChIP-seq, and assessment of ??-KG-dependent dioxygenase activity. It is also suitable for cell proliferation assays and functional rescue experiments using wild-type L2HGDH to confirm on-target effects. The model supports studies in cancer metabolism, epigenetic regulation, and functional genomics. For additional information or to discuss your specific experimental needs, please contact Ascent Research.
