The H2AX Knockout 786-O Polyclonal Cells are a CRISPR/Cas9-edited polyclonal population with targeted disruption of the H2AX gene, originating from the 786-O renal cell adenocarcinoma line. This gene-edited pool consists of a heterogeneous mixture of cells carrying diverse editing events, providing a loss-of-function model for investigating DNA double-strand break repair without clonal selection.
The parental 786-O cell line is a clear cell renal cell carcinoma-derived epithelial model extensively employed in cancer research. It harbors a characteristic VHL gene mutation resulting in constitutive stabilization of hypoxia-inducible factors and aberrant activation of mTOR signaling, thereby recapitulating key molecular features of human renal tumors. This genetic background makes 786-O an ideal host for dissecting DNA damage response mechanisms in the context of kidney cancer.
H2AX encodes a histone H2A variant that functions as a sentinel for DNA double-strand breaks. Upon genotoxic insult, the kinases ATM, ATR, and DNA-PKcs phosphorylate H2AX at Ser139 to produce ??H2AX, which rapidly forms nuclear foci at damage sites. ??H2AX serves as a molecular scaffold, recruiting the mediator protein MDC1, which in turn interacts with 53BP1 and the MRE11-RAD50-NBS1 complex, while also promoting BRCA1 accumulation. These complexes orchestrate homologous recombination and non-homologous end joining repair pathways, while ATM/ATR propagating signals through Chk1 enforce cell cycle checkpoints to prevent replication of damaged DNA.
In 786-O cells, abrogation of H2AX function disrupts the integrity of the DNA damage response, potentially exacerbating genomic instability. This model is particularly relevant given the high incidence of DNA repair defects and therapeutic resistance in renal cell carcinoma. By eliminating H2AX, researchers can explore how DNA repair proficiency influences cellular sensitivity to radiation and chemotherapeutic agents such as cisplatin and etoposide, which are commonly used in oncology and are known to generate double-strand breaks. The polyclonal nature of the knockout also mimics the genetic heterogeneity observed in tumor populations.
This knockout cell pool supports a broad range of experimental approaches, including western blotting and immunofluorescence microscopy for ??H2AX detection, clonogenic survival assays following ionizing radiation or drug exposure, comet assays to quantify DNA breakage, flow cytometry for cell cycle distribution profiling, and co-immunoprecipitation to examine repair complex assembly. It is suited for drug screening of genotoxic compounds, mechanistic studies of ATM/ATR signaling, and biomarker discovery for DNA repair deficiencies. For further information or technical support, please contact Ascent Research.
