CRISPR/Cas9-mediated gene disruption has been employed to generate a polyclonal JAG1 knockout cell population in the 786-O human renal cell carcinoma line. This polyclonal knockout product consists of a heterogeneous pool of cells harboring targeted disruptions in the JAG1 locus, enabling loss-of-function analysis without the clonal selection bias inherent in monoclonal isolates. The knockout cell pool provides a robust platform for studying JAG1-dependent signaling and tumorigenic processes in a VHL-mutant clear cell renal adenocarcinoma background.
The 786-O cell line originates from a primary clear cell renal adenocarcinoma and carries a well-characterized VHL tumor suppressor gene mutation, resulting in constitutive stabilization of hypoxia-inducible factors and aberrant angiogenic signaling. As epithelial-derived cancer cells, 786-O cells exhibit classic features of renal cell carcinoma, including anchorage-independent growth and high tumorigenic potential in xenograft models. This genetic and phenotypic context renders them highly relevant for investigating the interplay between VHL loss and Notch pathway activation in kidney cancer.
JAG1 encodes a transmembrane ligand that engages Notch receptors (NOTCH1?C4) to trigger sequential proteolytic cleavages by ADAM10/ADAM17 metalloproteases and the ??-secretase complex, releasing the Notch intracellular domain (NICD). NICD translocates to the nucleus, where it forms a transcriptional activation complex with RBPJ and MAML1, driving expression of target genes such as HES1, HEY1, MYC, and CCND1. JAG1 also crosstalks with other signaling modules, including PI3K/AKT, NF-??B, and Wnt/??-catenin pathways, and its activity is modulated by interacting partners like DLL1, DLL4, JAG2, NUMB, and E3 ubiquitin ligases MIB1 and DTX1 that regulate ligand endocytosis and signaling strength.
In 786-O cells, JAG1-mediated Notch activation contributes to oncogenic processes such as epithelial?mesenchymal transition (EMT), angiogenesis, and cell proliferation. JAG1 knockout in this VHL-mutant background disrupts the Notch-dependent transcriptional program, thereby attenuating the expression of prometastatic factors like SNAI1 and repressing the endothelial growth factor VEGFA, a key angiogenic driver in clear cell renal carcinoma. This knockout model thus recapitulates critical aspects of JAG1 loss in a renal cancer context and enables dissection of Notch pathway contributions independent of VHL modulation.
This polyclonal JAG1 knockout cell product supports a broad range of functional studies, including Notch signaling dissection via Western blotting for JAG1, NOTCH1, and HES1, RT-qPCR profiling of HES1, HEY1, and MYC, and immunofluorescence localization of NICD. The cells are well suited for EMT analysis by Transwell migration and invasion assays, apoptosis assessment by Annexin V staining, and in vivo xenograft tumor growth evaluation. Co-immunoprecipitation can be employed to probe JAG1?CNOTCH1 interactions, while Notch transcriptional reporter assays quantify pathway activity. These applications are central to investigations of renal cancer biology, drug resistance mechanisms, and tumor microenvironment interactions. For additional information or technical support, please contact Ascent Research.
