The AASS Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the near-haploid HAP1 cell line, carrying targeted disruption of the alpha-aminoadipic semialdehyde synthase (AASS) gene. This model provides a heterogeneous loss-of-function system for studying lysine catabolism, leveraging the HAP1 background favored for haploid genetic screening. The AASS gene encodes the bifunctional enzyme that initiates the saccharopine pathway of lysine degradation, making it a critical node in amino acid metabolism.
The HAP1 cell line, originating from a chronic myeloid leukemia patient, displays adherent fibroblast-like morphology and a near-haploid karyotype, facilitating robust CRISPR genome editing and functional genomics studies. It retains myeloid lineage traits and is extensively used for haploid genetic screening, drug target validation, and signaling pathway analysis, with proven utility in metabolic pathway investigations such as amino acid breakdown.
AASS functions as a homodimeric enzyme catalyzing the condensation of lysine with ??-ketoglutarate to form saccharopine, followed by hydrolysis to alpha-aminoadipic semialdehyde and glutamate. Its activity is regulated by nutritional lysine availability, mTORC1 signaling that couples amino acid sensing to cellular growth, and transcriptional control by the coactivator PPARGC1A. Downstream, the products alpha-aminoadipic semialdehyde and glutamate influence the NADH/NAD+ ratio and mitochondrial protein glutarylation. Disruption of AASS blocks these reactions, impairing lysine catabolic flux and leading to lysine accumulation and mitochondrial saccharopine metabolism imbalance, which are hallmarks of hyperlysinemia-related metabolic dysregulation.
In the HAP1 near-haploid context, AASS knockout offers a simplified genetic background for studying dose-dependent effects and synthetic lethal interactions relevant to lysine metabolism. This model is particularly pertinent for hyperlysinemia and lysine intolerance research, conditions linked to developmental delay and metabolic imbalance. The myeloid lineage of HAP1 also allows exploration of amino acid metabolism in hematological contexts, although AASS is not lineage-restricted, making the system broadly useful for metabolic disease investigation.
Typical applications include AASS enzyme activity assays to confirm loss of function, HPLC-based saccharopine measurement to monitor substrate accumulation, LC-MS lysine quantification for metabolic profiling, and Western blotting or RT-qPCR to assess pathway component expression. Viability assays under lysine restriction or supplementation, metabolic flux analysis using stable isotope tracing, and CRISPR editing validation are readily performed. The polyclonal population also enables haploid genetic interaction screens to identify modulators of the saccharopine pathway and drug sensitivity studies. For further details, please contact Ascent Research.
