AHNAK Knockout HEK293T Polyclonal Cells from Ascent Research provide a CRISPR/Cas9-edited polyclonal knockout cell population designed for studying AHNAK (neuroblast differentiation-associated protein AHNAK) function. This loss-of-function model enables disruption of target gene expression without pre-defined clone isolation, offering a genetically heterogeneous population suitable for pooled functional studies.
The knockout is performed in the HEK293T host cell line, a widely used human embryonic kidney epithelial model stably expressing the SV40 large T antigen. HEK293T cells exhibit high transfection efficiency and support episomal replication of plasmids containing the SV40 origin, making them an ideal chassis for transient and stable genetic manipulation. Their epithelial origin and robust growth characteristics provide a reliable platform for investigating scaffold protein functions in cell adhesion and signaling.
AHNAK functions as a large scaffolding protein that integrates calcium signaling with actin cytoskeleton remodeling. It is activated downstream of TGF-beta receptor and calcium influx, and regulated by PKC phosphorylation. AHNAK interacts with the annexin A2?CS100A10 complex and F-actin, positioning it at membrane-cytoskeleton interfaces. This scaffold coordinates the activation of RhoA and Rac1 small GTPases to control actin dynamics and cell adhesion, while also modulating SMAD2/3-dependent TGF-beta transcriptional responses. Additionally, AHNAK associates with dystrophin, calpain, and Hsp90, linking it to membrane integrity and stress responses. Through these interactions, AHNAK influences L-type calcium channel activity and calcium-dependent signaling cascades involving CaMKII and calcineurin.
In HEK293T cells, AHNAK-dependent pathways regulate epithelial cell adherence and migration, processes recapitulated in vitro using standard scratch-wound and transwell assays. The polyclonal knockout population enables researchers to examine heterogeneous loss-of-function effects, revealing the spectrum of cellular responses dependent on AHNAK expression. Given the extensive crosstalk between AHNAK and TGF-beta, calcium, and cytoskeletal pathways, this model is particularly valuable for dissecting signaling networks that govern epithelial-to-mesenchymal transition and cancer cell dissemination.
Applications include investigating the role of AHNAK in breast cancer, hepatocellular carcinoma, and melanoma progression, where its scaffolding functions influence invasion and metastasis. The knockout cells also serve as a platform for screening small-molecule modulators of AHNAK-anchored signaling nodes. Researchers can validate protein interactions by co-immunoprecipitation of AHNAK with annexin A2, S100A10, or dystrophin, and assess TGF-beta pathway activity using SMAD-responsive luciferase reporters. Functional assays such as calcium imaging and F-actin staining further elucidate the scaffold??s impact on second-messenger dynamics and cytoskeletal organization. For inquiries about batch-specific knockout efficiency or custom modifications, please contact Ascent Research.
