The ATXN2 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population designed for comprehensive loss-of-function analysis of the ATXN2 gene. Derived from the HEK293T host line, this heterogeneous pool carries targeted gene disruption at the ATXN2 locus, resulting in ablation of ATXN2 protein expression. The polyclonal format minimizes clonal selection artifacts and provides a model representative of the diverse genetic background encountered in population-level studies.
HEK293T is a human embryonic kidney epithelial cell line immortalized by adenovirus type 5 E1A/E1B gene expression and stably expressing the SV40 large T-antigen. This genetic modification endows the cells with high transfection efficiency and the capacity for episomal replication of SV40 origin-containing plasmids, making them a gold standard for recombinant protein production, viral packaging, and lentivirus generation. The ATXN2 knockout derivative maintains these advantageous traits, thereby enabling transient or stable complementation with wild-type or mutant ATXN2 in rescue experiments.
ATXN2 encodes a conserved RNA-binding protein that shuttles between polysomes, stress granules, and processing bodies (P-bodies) to orchestrate mRNA translation, stability, and subcellular localization. ATXN2 is activated by diverse cellular stress stimuli, including oxidative stress and heat shock, downstream of mTORC1 and eIF2?? kinase signaling. It directly interacts with poly(A)-binding protein (PABPC1), TDP-43, G3BP1, TIA-1, DDX6, LSM12, and the IP3 receptor (IP3R), forming ribonucleoprotein complexes that control the fate of target mRNAs such as TARDBP and ATXN2 itself. In the absence of ATXN2, stress granule assembly is impaired, and the mRNA decay machinery involving UPF1 and XRN1 is dysregulated. Pathogenic expansions of the polyglutamine tract in ATXN2 drive protein aggregation and are implicated in spinocerebellar ataxia type 2, amyotrophic lateral sclerosis, and Parkinson??s disease, underscoring the clinical importance of this model.
The HEK293T background offers a tractable system for dissecting ATXN2-dependent pathways. The knockout cells are particularly valuable for studying the recruitment of TDP-43 to stress granules and for evaluating the interplay between ATXN2 and IP3R-mediated calcium signaling. The high protein expression capacity of HEK293T facilitates transient overexpression of ATXN2 variants, enabling structure?Cfunction analyses and co-immunoprecipitation of intact ribonucleoprotein assemblies. This model also supports quantitative assessment of autophagy and protein aggregation phenotypes.
Typical experimental applications include immunofluorescence-based monitoring of G3BP1 and TIA-1 stress granule markers, western blotting for ATXN2 and TDP-43, co-immunoprecipitation to map protein?Cprotein interactions, RT-qPCR to measure mRNA stability of ATXN2 targets, and calcium imaging to probe IP3R signaling. Additionally, these cells are suited for compound screening using cell viability or protein aggregation readouts in neurodegenerative disease research. For further technical assistance and detailed protocols, please contact Ascent Research.
