The ATIC Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population with targeted disruption of the ATIC gene in the HAP1 human near-haploid cell line. This polyclonal format encompasses a heterogeneous array of loss-of-function alleles, avoiding clonal selection artifacts and enabling robust functional analysis of de novo purine biosynthesis and its downstream effects.
The HAP1 cell line, derived from KBM-7 chronic myeloid leukemia cells, exhibits a near-haploid fibroblast-like phenotype. Its haploid genome simplifies genetic knockout studies by eliminating the complexity of biallelic targeting, making it highly suited for CRISPR-based screening. HAP1 cells demonstrate rapid growth and karyotypic stability, supporting reproducible biochemical and pharmacological assays in the context of nucleotide metabolism and cell signaling.
ATIC encodes a bifunctional enzyme catalyzing the final two steps of de novo purine biosynthesis: AICAR transformylase and IMP cyclohydrolase, which convert AICAR to IMP. It assembles into the purinosome complex with interacting factors GART, PAICS, and ADSL. Upstream regulators include transcription factors MYC and E2F1, and the substrate PRPP, while folate status influences one-carbon donor availability. ATIC knockout results in accumulation of AICAR, which allosterically activates AMPK, a central energy sensor. Concurrently, downstream metabolites IMP, AMP, and GMP are depleted, disrupting nucleotide pools critical for DNA and RNA synthesis and compromising cell cycle progression.
Within the HAP1 genetic background, ATIC knockout offers a precise model to dissect the interplay between purine metabolism and AMPK signaling. The haploid state ensures direct genotype-phenotype correlation, free from confounding wild-type alleles. This system is particularly relevant for modeling AICA-ribosiduria, a rare neurometabolic disorder, and for exploring cancer cell metabolism, where enhanced de novo purine synthesis supports proliferation. Accumulation of AICAR and consequent AMPK activation can be quantitatively tracked, providing insights into metabolic stress adaptation.
Applications include high-throughput screening of compounds that modulate purine biosynthesis, detailed AMPK signaling analysis via Western blotting and phospho-specific flow cytometry, and quantitative nucleotide pool profiling by LC-MS. Cell proliferation and viability assays can assess growth in purine-depleted conditions. The model also facilitates drug target validation for anticancer agents and functional investigations using RT-qPCR to measure purinosome gene expression. For further technical details, please contact Ascent Research.
