The HDHD3 Knockout HeLa Polyclonal Cells provide a CRISPR/Cas9-edited polyclonal knockout cell population in which the gene encoding pseudouridine-5′-phosphatase (HDHD3) has been disrupted. This polyclonal pool enables loss-of-function studies without clonal selection, maintaining genetic heterogeneity while abrogating HDHD3 protein expression across the population. The product is designed for researchers investigating pseudouridine metabolism, nucleotide salvage, and cancer cell metabolic reprogramming.
These knockout cells are engineered in the well-established HeLa host cell line, originally derived from a cervical adenocarcinoma. HeLa cells are HPV18-positive, expressing the E6 and E7 oncoproteins that inactivate the tumor suppressors p53 and Rb, respectively. The line exhibits a hypertriploid karyotype with numerous chromosomal abnormalities, providing a robust model for cancer biology, virology, and protein expression studies. This background makes the HDHD3 knockout particularly relevant for exploring oncogene-driven metabolic alterations.
HDHD3 functions as a pseudouridine-5′-phosphatase that catalyzes dephosphorylation of pseudouridine 5′-phosphate to pseudouridine, a critical step in pyrimidine catabolism and nucleotide salvage. Within this pathway, HDHD3 acts upstream of pseudouridine kinase and pseudouridine-5′-phosphate glycosidase, linking to uridine phosphorylase. Its activity is regulated by substrate availability and is putatively modulated by transcription factors MYC and HIF1A. Downstream, knockout of HDHD3 leads to reduced pseudouridine dephosphorylation, accumulation of pseudouridine 5′-phosphate, and altered nucleotide pool balance, potentially affecting tRNA and rRNA modification dynamics. Interacting factors include nucleoside kinases and phosphatases that coordinate nucleotide homeostasis.
In the HeLa context, disruption of HDHD3 intersects with the cell’s HPV-driven metabolic reprogramming. The loss of pseudouridine-5′-phosphatase activity is expected to perturb nucleotide pools and RNA modification recycling, which may impact cell proliferation and stress responses. This model is especially informative for cancer metabolism research, as HeLa cells rely on robust nucleotide metabolism to sustain rapid division. The knockout thus offers a platform to dissect how pyrimidine catabolism contributes to oncogenic fitness and to identify vulnerabilities in pseudouridine degradation pathways.
This polyclonal HDHD3 knockout cell population is suitable for a range of applications, including functional dissection of pseudouridine metabolism, investigation of nucleotide salvage pathways, and screening for metabolic dependencies in cancer cells. Recommended assays include Western blotting and RT-qPCR for HDHD3 loss verification, pseudouridine 5′-phosphatase activity measurements, LC-MS-based metabolite profiling, nucleotide pool quantification, and cell proliferation analyses. Researchers can leverage this model to uncover novel roles of HDHD3 in RNA modification dynamics and tumor cell fitness. For further details or to explore custom applications, please contact Ascent Research.
