The ALDH4A1 Knockout HT29 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal knockout cell population engineered for loss-of-function analysis of the ALDH4A1 gene in a human colorectal adenocarcinoma background. This product provides a heterogeneous pool of HT29 cells with targeted disruption of ALDH4A1 via CRISPR/Cas9-mediated gene editing, enabling researchers to study the collective effects of ALDH4A1 ablation without clonal selection. The knockout model serves as a versatile tool for investigating proline metabolism, mitochondrial function, and amino acid catabolism in cancer biology.
The HT29 parental cell line is derived from a human colorectal adenocarcinoma and exhibits an epithelial morphology characteristic of intestinal epithelial cells. As a widely used model in colorectal cancer research, HT29 cells retain key features of the intestinal epithelium and are amenable to metabolic, signaling, and functional assays. This adherent cell line grows readily in standard culture conditions and is commonly employed to study tumor cell proliferation, migration, drug sensitivity, and metabolic adaptations.
ALDH4A1 encodes a mitochondrial NAD-dependent dehydrogenase that irreversibly converts delta-1-pyrroline-5-carboxylate (P5C) to glutamate, a critical step in the degradation of proline and arginine. This reaction directly couples amino acid catabolism to the tricarboxylic acid (TCA) cycle, as glutamate can be deaminated by GLUD1 (glutamate dehydrogenase) to ??-ketoglutarate, thereby fueling oxidative phosphorylation and ATP production. ALDH4A1 activity is regulated by the availability of proline and arginine and by transcriptional programs governing amino acid metabolism. The enzyme interacts with NAD+ as a cofactor and functionally cooperates with other mitochondrial dehydrogenases and TCA cycle enzymes to maintain cellular energy homeostasis.
In HT29 colorectal cancer cells, disruption of ALDH4A1 is predicted to impair the conversion of P5C to glutamate, potentially leading to accumulation of P5C and altered mitochondrial respiration. This metabolic blockade may affect the TCA cycle flux, reduce ATP output, and shift the metabolic profile of the cells, which is particularly relevant in the context of cancer cell metabolism where proline catabolism supports bioenergetic and biosynthetic demands. The model thus offers a platform to dissect the contribution of proline metabolism to tumor growth, survival under nutrient stress, and the development of chemoresistance.
This knockout cell population is ideally suited for a range of advanced research applications, including metabolic profiling to trace carbon flux from proline into the TCA cycle, Seahorse-based measurements of oxygen consumption and mitochondrial function, and Western blot or RT-qPCR analysis of TCA cycle enzymes and metabolic regulators. It can also be employed in cell proliferation assays, migration studies, and drug sensitivity screens to evaluate how ALDH4A1 loss influences cancer cell behavior. By linking amino acid degradation to energy metabolism, the model enables investigation of metabolic vulnerabilities in colorectal cancer and may inform therapeutic strategies targeting metabolic pathways. For further information, contact Ascent Research.
