The KBTBD2 Knockout HEK293T Polyclonal Cells product provides a CRISPR/Cas9-edited polyclonal population of HEK293T cells with targeted disruption of the KBTBD2 gene. This knockout model enables loss-of-function studies of KBTBD2, a substrate-specific adaptor for the Cullin3-RING E3 ubiquitin ligase complex. The polyclonal nature ensures a mixed genetic background that may capture diverse mutational events, suitable for population-level analyses. This tool is designed for researchers investigating the ubiquitin-proteasome system and insulin signaling regulation.
HEK293T cells are derived from human embryonic kidney epithelial cells and are immortalized by the SV40 large T antigen, which promotes episomal replication of transfected plasmids and high-level protein expression. These cells are widely employed for protein overexpression, viral production, and signal transduction studies due to their ease of culture and transfectability. The epithelial origin and kidney lineage provide a physiologically relevant platform for studying metabolic pathways, although the transformed nature may introduce caveats in differentiation-related contexts.
KBTBD2 functions as a substrate recognition component of the CUL3-RBX1 E3 ubiquitin ligase complex, directing the polyubiquitination of insulin receptor substrate 1 (IRS1) for proteasomal degradation. This activity is modulated by upstream signals including insulin stimulation and the NEDD8 conjugation system, which activates cullin-RING ligases. Through targeted degradation of IRS1, KBTBD2 negatively regulates downstream insulin signaling, attenuating AKT phosphorylation and impairing glucose uptake. Additionally, KBTBD2 interacts with CUL3 and RBX1, and its expression may be influenced by myogenic differentiation cues, linking it to metabolic and developmental processes.
In HEK293T cells, the knockout of KBTBD2 is expected to stabilize IRS1 protein levels and potentiate insulin-induced signaling, making this model particularly valuable for dissecting feedback mechanisms within the insulin pathway. Given the cells?? kidney epithelial origin, this system also permits investigation of renal glucose handling and its dysregulation in metabolic diseases. However, because HEK293T cells are not insulin target tissues like muscle or adipose, results should be interpreted with consideration of cell type-specific signaling networks. The polyclonal knockout pool allows for the study of gene function without clonal artifacts, reflecting a broader spectrum of cellular responses.
This knockout cell model supports a range of experimental applications, including Western blot analysis of IRS1 and phospho-AKT to assess insulin sensitivity, co-immunoprecipitation to confirm disrupted KBTBD2-CUL3 interactions, and in-cell ubiquitination assays to monitor IRS1 degradation dynamics. It is also suitable for insulin-stimulated glucose uptake measurements and RT-qPCR profiling of metabolic gene expression. Researchers in diabetes, obesity, and ubiquitin biology will find this tool invaluable for dissecting the molecular determinants of insulin resistance and metabolic syndrome. For further information or technical support, please contact Ascent Research.
