The ALDH18A1 Knockout HeLa Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population engineered from the HeLa cell line for loss-of-function analysis of ALDH18A1, the gene encoding delta-1-pyrroline-5-carboxylate synthase (P5CS). This polyclonal product offers a heterogeneous mixture of cells carrying targeted disruptions at the ALDH18A1 locus, facilitating pooled investigations of gene function without the limitations of single-cell-derived clones. The model serves as a versatile reagent for studying the immediate and downstream metabolic consequences of P5CS inactivation.
HeLa cells are an HPV18-positive cervical adenocarcinoma epithelial line in which viral oncoproteins E6 and E7 suppress p53 and RB, leading to a highly aneuploid and genomically unstable phenotype. This widely used immortalized background is particularly relevant for cancer metabolism research, as p53 loss and RB inactivation influence nutrient sensing and stress responses, making it an appropriate host to examine the impact of ALDH18A1 knockout on amino acid metabolic pathways.
ALDH18A1 encodes the bifunctional enzyme P5CS, which catalyzes the ATP- and NADPH-dependent conversion of glutamate to pyrroline-5-carboxylate (P5C), the rate-limiting step in proline biosynthesis. P5C is reduced to proline by PYCR1/2 or transaminated to ornithine, feeding into arginine synthesis and the urea cycle. ALDH18A1 transcription is driven by c-MYC, ATF4, and mTORC1, and repressed by p53, linking its activity to growth signaling and nutrient status. Proline, ornithine, and arginine serve as precursors for collagen, polyamines, and nitric oxide, respectively, underscoring the central role of P5CS in coordinating biosynthetic and signaling outputs from glutamate metabolism.
In the HeLa cervical cancer model, proline metabolism supports rapid proliferation and stress adaptation; P5CS-dependent proline synthesis may protect these cells against oxidative damage and nutrient deprivation. ALDH18A1 disruption is thus expected to impair proline availability, rendering cells susceptible to metabolic stress, ferroptosis, or amino acid limitation. This knockout system offers a powerful tool to investigate how proline biosynthesis interplays with oncogenic signaling in p53- and RB-deficient tumors and to explore ALDH18A1 as a metabolic vulnerability.
This ALDH18A1 knockout HeLa polyclonal population is suitable for investigating proline and arginine metabolism in cancer through metabolic flux analyses using isotopically labeled glutamate, cell proliferation and clonogenic survival assays under nutrient deprivation, and sensitivity profiling with metabolic inhibitors. Complementary validation techniques include western blotting, RT-qPCR, and mass spectrometry-based quantification of proline, ornithine, and arginine. Transcriptomic adaptation can be monitored by RNA-seq, while amino acid starvation sensitivity assays directly assess metabolic resilience. For further details or to request a quote, please contact Ascent Research.
