The DNASE1L1 knockout HT29 polyclonal cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 human colorectal adenocarcinoma cell line. They harbor targeted disruption of the DNASE1L1 gene, leading to loss of deoxyribonuclease 1-like 1 protein. As a polyclonal pool, they contain diverse loss-of-function alleles, offering a robust model without clonal selection bias. The knockout was generated via CRISPR/Cas9-mediated gene editing, introducing targeted disruptions within the DNASE1L1 locus to inactivate gene function. This loss-of-function tool is designed for advanced research into apoptosis, DNA degradation, and related pathways.
HT29 cells are an established human colorectal adenocarcinoma line, isolated from a primary tumor of a 44-year-old female. They display adherent epithelial morphology and retain intestinal epithelial features, including polarized monolayer formation and mucin secretion. Widely used in cancer research, HT29 cells model colorectal cancer biology, drug resistance, and epithelial barrier function. Their genetic background includes mutations in APC and p53, providing a clinically relevant system for studying tumorigenesis and therapeutic responses. This context allows meaningful investigation of DNASE1L1 loss in colorectal malignancy.
The DNASE1L1 gene encodes a Ca2+/Mg2+-dependent endonuclease that digests chromatin DNA into nucleosomal fragments during apoptosis. It is regulated upstream by TNF-alpha and p53, and functions downstream of caspase-3. DNASE1L1 interacts with DNA, histones, and actin, and cooperates with nucleases such as CAD/DFF40 and EndoG, while ICAD modulates activity. This nuclease drives efficient chromatin degradation, facilitating apoptotic cell disassembly and preventing immunogenic DNA release. The process is critical for lysosomal DNA degradation and innate immune tolerance, linking DNASE1L1 to apoptosis, necrosis, and inflammatory signaling.
In HT29 colorectal cancer cells, DNASE1L1 knockout disrupts apoptotic DNA degradation, exacerbating defects in apoptosis and DNA repair common in colorectal tumors. Undegraded DNA accumulates and acts as a DAMP, promoting inflammation and influencing the tumor microenvironment. This model helps dissect how impaired DNA clearance shifts cell death from apoptosis to necrosis and modulates immune activation. Additionally, it offers insight into intestinal epithelial barrier dysfunction and inflammatory bowel disease. Thus, it provides a valuable system for investigating DNA metabolism, cell death, and colorectal cancer progression.
These DNASE1L1 knockout HT29 polyclonal cells support diverse applications. They enable mechanistic studies of caspase-dependent DNA fragmentation via TUNEL and DNA laddering, and apoptotic quantification by Annexin V flow cytometry. The model facilitates high-throughput screening of colorectal cancer drugs targeting apoptosis, and studies of intestinal homeostasis under genotoxic stress. RT-qPCR and Western blotting allow signaling pathway analysis. Moreover, they help explore crosstalk between apoptosis and innate immunity, including how defective DNA degradation triggers cytokine release. For further information, contact Ascent Research.
