This product provides a CRISPR/Cas9-edited polyclonal knockout cell population targeting the EIF5B gene in the HeLa human cell line. The polyclonal knockout cells are generated through CRISPR/Cas9-mediated gene disruption, resulting in a heterogeneous pool of EIF5B-deficient HeLa cells. This loss-of-function model is an essential tool for dissecting the role of EIF5B in translation initiation and its broader impact on cellular processes.
HeLa cells are an immortalized human cervical adenocarcinoma line originally derived from Henrietta Lacks, positive for human papillomavirus type 18 (HPV18). These epithelial cells are one of the most extensively used models in cancer biology, owing to their robust growth, high transfection efficiency, and well-characterized signaling networks. The HeLa background provides a physiologically relevant context for studying translation regulation in a cancer setting, particularly given the reliance of tumor cells on enhanced protein synthesis for proliferation and survival.
EIF5B encodes a GTPase that catalyzes the joining of the 40S and 60S ribosomal subunits to form the 80S initiation complex, a rate-limiting step in cap-dependent translation. Mechanistically, EIF5B interacts with eIF1A, eIF5, the 40S and 60S subunits, GTP, and initiator tRNA to mediate subunit joining. GTP hydrolysis by EIF5B triggers conformational changes that release initiation factors and commit the ribosome to elongation. Upstream, EIF5B activity is controlled by the mTORC1 pathway and eIF2 kinases, integrating signals from growth factors and amino acid availability. Downstream, functional EIF5B drives global protein synthesis by facilitating 80S ribosome assembly, placing it at a convergence point of the mTOR signaling and integrated stress response networks.
In HeLa cells, which exhibit high translational output driven by HPV oncoproteins and mTOR hyperactivity, disruption of EIF5B significantly impairs the formation of translation-competent 80S ribosomes. This leads to attenuated global protein synthesis and can activate stress-responsive pathways, making this knockout model valuable for investigating how translational dysregulation contributes to cancer cell viability. Additionally, because many viruses hijack the host translation machinery via interactions with initiation factors including eIF5B, these cells offer a platform for dissecting viral replication mechanisms and evaluating host-directed antiviral strategies.
Researchers can employ this polyclonal EIF5B-knockout HeLa population in a wide range of assays, including polysome profiling and ribosome profiling to monitor translation initiation defects, dual-luciferase reporter assays to quantify cap-dependent translation, co-immunoprecipitation to probe interactions with ribosomal subunits and initiation factors, and GTPase activity measurements to assess the functional impact of EIF5B disruption. Moreover, western blotting and RT-qPCR enable verification of knockout efficiency and downstream effects on protein expression. This product is suited for applications in cancer biology, viral replication studies, and discovery of antiproliferative agents targeting the protein synthesis machinery. For additional technical details or custom requests, please contact Ascent Research.
