The AFF2 Knockout HeLa Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the human HeLa cervical adenocarcinoma cell line. These cells carry a targeted disruption of the AFF2 gene, resulting in a loss-of-function model that ablates expression of the encoded scaffold protein for the super elongation complex (SEC). The polyclonal format comprises a heterogeneous mixture of edited cells, each harboring distinct CRISPR/Cas9-mediated genomic alterations at the AFF2 locus, providing robust population-level analyses suitable for downstream applications.
The parental HeLa cell line is an adherent, HPV18-positive cervical adenocarcinoma epithelial model extensively used in cancer biology. The integrated HPV18 E6 and E7 oncoproteins constitutively inactivate the p53 and retinoblastoma (Rb) tumor suppressors, respectively, generating a background of unchecked cell cycle progression and transcriptional dysregulation. This well-characterized environment permits precise interrogation of how AFF2-dependent transcriptional mechanisms interface with oncogenic signaling pathways.
Mechanistically, AFF2 scaffolds the super elongation complex (SEC) and recruits P-TEFb (CDK9/CCNT1) to RNA polymerase II, releasing promoter-proximal pausing and enabling transcription elongation. It also binds ??-catenin (CTNNB1) to co-activate Wnt target genes, including MYC and CCND1. The SEC incorporates ELL, MLLT1/ENL, and MLLT3/AF9. Upstream regulation involves Wnt ligands (e.g., WNT3A) and CTNNB1, as well as CDK9 kinase activity, while downstream targets encompass the neurodevelopmental gene GRIN2B. Thus, AFF2 integrates elongation control with Wnt/??-catenin signaling.
In the HeLa context, AFF2 knockout disrupts SEC assembly and Wnt/??-catenin transcriptional responses, providing a system to dissect the interplay between transcriptional elongation and oncogenic signaling. This model is valuable for studying how loss of AFF2 impacts proliferation, apoptosis, and the expression of Wnt targets in a transformed epithelial background. Additionally, it serves as a tractable platform to investigate conserved molecular mechanisms underlying fragile X E syndrome and intellectual disability, despite the non-neuronal origin of HeLa cells.
Typical applications include ChIP-qPCR to measure Pol II occupancy, RNA-seq for transcriptome-wide elongation and splicing analysis, and co-immunoprecipitation to assess SEC integrity. Western blotting can validate downstream effector changes, while Wnt-responsive luciferase reporter assays quantify ??-catenin activity. Cell proliferation and apoptosis assays deliver phenotypic insights, and immunofluorescence microscopy tracks protein localization. For additional details or to inquire about custom formulations, please contact Ascent Research.
