The EHBP1 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout population derived from HEK293T human embryonic kidney cells. This loss-of-function model targets EHBP1, which encodes a scaffold protein linking endocytic trafficking to the actin cytoskeleton. The polyclonal pool is generated by transient delivery of CRISPR/Cas9 ribonucleoprotein complexes, creating a heterogeneous population with targeted gene disruption. This cost-effective format is ideal for bulk assays where clonal isolation is unnecessary.
HEK293T cells are a human embryonic kidney line immortalized with adenovirus 5 DNA and expressing SV40 large T antigen. They are highly transfectable and support robust protein expression, making them a versatile platform for signal transduction, viral production, and membrane trafficking studies. These cells express insulin receptor, PI3K, and Akt, thus providing a suitable background for analyzing insulin-regulated processes like GLUT4 translocation. Although not an insulin-responsive tissue, the ease of genetic manipulation enables detailed molecular dissection of trafficking and cytoskeletal dynamics.
EHBP1 (EH domain-binding protein 1) functions as an adaptor coupling endocytic vesicles to the cortical actin network, a process essential for insulin-stimulated GLUT4 translocation. Activation of the insulin receptor leads to PI3K and Akt signaling, which regulates EHBP1-mediated vesicle trafficking. EHBP1 interacts with Eps15 and Rab8a to coordinate actin polymerization and vesicle tethering. CRISPR/Cas9-mediated disruption abrogates these interactions, causing impaired actin remodeling and defective GLUT4 translocation, thereby attenuating glucose uptake. This mechanistic framework highlights EHBP1 as a key node linking insulin receptor activation to cytoskeletal reorganization and glucose homeostasis.
In the HEK293T background, EHBP1 knockout provides a simplified model to study GLUT4 trafficking and actin dynamics. Although these cells do not endogenously express GLUT4, ectopic expression allows reconstitution of insulin-responsive translocation. The loss-of-function model enables dissection of EHBP1’s role in endocytic recycling and actin assembly, independent of adipocyte or muscle physiology. This system is valuable for identifying interacting partners and signaling inputs that modulate EHBP1 activity.
This polyclonal knockout product is suitable for insulin signaling studies, glucose metabolism experiments, vesicle trafficking analysis, and actin cytoskeleton investigations. Representative assays include Western blotting for EHBP1 and phospho-Akt, immunofluorescence for GLUT4 translocation, glucose uptake measurements, co-immunoprecipitation of EHBP1 interactors, RT-qPCR for target gene disruption, and actin polymerization assays. The model can also be employed for drug screening to identify insulin sensitizers. For further information, please contact Ascent Research.
