The GTF2H2 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population targeting the GTF2H2 gene, which encodes the p44 subunit of the TFIIH complex. This heterogeneous pool of HEK293T cells features gene disruptions that ablate functional p44, providing a versatile loss-of-function model. The polyclonal format captures diverse editing outcomes without clonal selection bias. These cells are designed for investigating TFIIH-dependent transcription and DNA repair processes.
HEK293T is a human embryonic kidney epithelial cell line transformed with SV40 large T antigen, enabling efficient plasmid replication and protein expression. Its robust growth, high transfectability, and suitability for CRISPR editing make it an ideal chassis for gene knockout studies. The renal epithelial origin offers a physiologically relevant background for studying DNA damage responses and cell cycle control. This host line is widely employed in recombinant protein production, viral packaging, and functional genomics.
GTF2H2??s p44 subunit is integral to TFIIH, a complex essential for RNA polymerase II (Pol II) transcription initiation and nucleotide excision repair (NER). p44 directly interacts with TFIIH subunits p62, p52, p34, XPB, XPD, and the CDK7?CCyclin H?CMAT1 kinase module. Upstream, ATM and ATR kinases activate TFIIH following DNA damage. Subsequently, TFIIH phosphorylates the Pol II C-terminal domain and recruits repair factors XPA, XPG, and XPF-ERCC1, along with RPA, to excise UV-induced lesions. Thus, GTF2H2 orchestrates the coupling of transcription to genome repair.
Disruption of GTF2H2 in HEK293T cells impairs TFIIH assembly and function, leading to defective Pol II transcription and NER. The cells exhibit increased UV sensitivity, impaired RNA synthesis recovery after UV, and accumulation of DNA damage. This phenotype models human trichothiodystrophy, xeroderma pigmentosum, and Cockayne syndrome, disorders linked to TFIIH mutations. The model facilitates mechanistic studies of transcription-repair coupling and cancer predisposition. It also provides a platform for screening compounds that modulate DNA repair or transcriptional checkpoints.
These polyclonal knockout cells support a range of experimental approaches. Transcriptional activity can be assessed by RNA-seq, RT-qPCR, and ChIP-qPCR for Pol II occupancy. DNA repair efficiency is evaluated via UV clonogenic survival, comet assay, and RNA synthesis recovery. Protein interactions are analyzable through co-immunoprecipitation and immunofluorescence of TFIIH components. Applications include UV sensitization screening, chemotherapeutic testing, and functional dissection of transcription-coupled repair. For additional information, contact Ascent Research.
