The E2F4 Knockout 786-O Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout cell population designed to disrupt expression of the E2F4 transcription factor in the human clear cell renal cell carcinoma line 786-O. This heterogeneous loss-of-function model is generated through bulk gene disruption, providing a tool free from clonal biases for interrogating E2F4-dependent transcriptional regulation.
The 786-O host cell line originates from a primary renal cell adenocarcinoma and is widely used as a model of clear cell renal cell carcinoma (ccRCC). It harbors an inactivating mutation in the VHL tumor suppressor, leading to constitutive activation of hypoxia-inducible factor pathways characteristic of ccRCC. This epithelial background offers a clinically relevant context for studying cell cycle dysregulation in renal cancer.
E2F4 is a transcriptional repressor that binds E2F promoter elements as a heterodimer with TFDP1 or TFDP2. It recruits pocket proteins p130 (RBL2) and p107 (RBL1), along with co-repressors HDAC1 and SIN3A, to silence G1/S cell cycle genes such as CCNA2, CCNE1, CDK1, MYC, and BIRC5. E2F4 activity is regulated by CDK4/6 phosphorylation of pocket proteins, which relieves repression, and by upstream signals including TGF-beta and p16INK4a, which promote E2F4?Cp130 complex formation. E2F4 thus integrates growth-inhibitory cues to maintain quiescence.
In the VHL-mutant 786-O context, E2F4 knockout disrupts cell cycle checkpoints, derepressing E2F targets and potentially accelerating proliferation. This polyclonal pool allows investigation of how E2F4 loss cooperates with HIF dysregulation to drive oncogenic phenotypes and alter sensitivity to CDK4/6 inhibitors or other cell cycle-targeted agents.
Applications include ChIP-qPCR to assess E2F4 promoter occupancy, RT-qPCR and western blotting for target gene and protein expression (e.g., cyclin E, CDK2), flow cytometry for cell cycle profiling, and functional assays such as MTT proliferation and colony formation. The polyclonal nature reduces clonal artifacts, making it suitable for CRISPR screen validation, epigenetic studies, and drug sensitivity profiling. For further inquiries, contact Ascent Research.
