The DTD2 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HAP1 cell line, featuring targeted disruption of the DTD2 gene. This loss-of-function model enables investigation of D-aminoacyl-tRNA deacylase function in a near-haploid human background. The polyclonal format provides a heterogeneous population of edited cells, reflecting varied indel patterns induced by CRISPR/Cas9, without clonal selection, offering a robust tool for studying gene function.
HAP1 is a near-haploid human cell line derived from the chronic myeloid leukemia (CML) cell line KBM-7, exhibiting adherent, fibroblast-like morphology and male origin. Its haploid nature makes it exceptionally suited for genetic screens and functional genomics, as the presence of a single copy of autosomal genes simplifies genotype-phenotype correlations. The cells proliferate in adherent culture and maintain stable haploidy under standard conditions, providing a controlled platform for interrogating gene function.
DTD2 (D-aminoacyl-tRNA deacylase 2) is a key translation quality control enzyme that hydrolyzes mischarged D-aminoacyl-tRNAs, preventing the incorporation of D-amino acids into nascent polypeptides. It forms interactions with ribosomes and aminoacyl-tRNA synthetases, and is implicated in mitochondrial protein synthesis through association with mitochondrial ribosomal proteins. Upstream signaling likely involves general translation regulators such as mTOR, while its activity maintains translational fidelity, thereby reducing proteotoxic stress caused by aberrant D-amino acid-containing proteins. Disruption of DTD2 may lead to accumulation of mischarged tRNAs and trigger mitochondrial dysfunction, highlighting its role in protein quality control.
In the HAP1 background, DTD2 knockout offers a clean genetic system to dissect the consequences of impaired translation fidelity in leukemia-derived cells. The near-haploid karyotype eliminates confounding effects from wild-type alleles, simplifying interpretation of knockout phenotypes. Studies can focus on mitochondrial stress responses, D-amino acid misincorporation, and downstream effects on cell viability. Given the CML origin of HAP1, this model also holds relevance for leukemia biology, enabling exploration of how translational quality control impacts cancer cell fitness.
These knockout cells are suitable for applications such as haploid genetic screens to identify modifiers of translational fidelity, mitochondrial translation studies, and functional assays assessing DTD2’s role in protein synthesis. Representative assays include western blotting for DTD2 and translation markers, puromycin incorporation to monitor global translation, tRNA charging assays, mitochondrial isolation and functional assessments, and flow cytometry for apoptosis or proliferation. The polyclonal population is ideal for pooled loss-of-function experiments and subsequent validation. For additional information or custom configurations, please contact Ascent Research.
