The HLA-DRA Knockout A-549 Polyclonal Cells comprise a CRISPR/Cas9-edited polyclonal knockout population of A-549 cells in which the HLA-DRA gene is disrupted to create a loss-of-function model for the MHC class II alpha chain. This heterogeneous pool of edited cells avoids clonal selection bias, enabling population-level analysis of HLA-DRA function. Validation using western blotting and flow cytometry confirms reduced HLA-DR protein expression.
A-549 is a human non-small cell lung cancer (NSCLC) adenocarcinoma cell line isolated from a male patient, widely used as a model of alveolar type II epithelial cells. This line is a cornerstone in respiratory research, drug metabolism, and NSCLC biology, retaining features like surfactant production and inducible MHC class II expression. IFN-gamma upregulates HLA-DRA in A-549 cells, making them a relevant system to study antigen presentation in the lung epithelium.
HLA-DRA encodes the invariant alpha chain of the HLA-DR heterodimer, which pairs with the beta chain (HLA-DRB) to present exogenous peptides to CD4+ T cells. Constitutive expression is limited, but transcription is tightly controlled by CIITA, which is activated by IFNG signaling, and the RFX complex (RFX5, RFXAP, RFXANK) along with NFYC and CREB1. The HLA-DR complex associates with the invariant chain (CD74) for trafficking to endosomal compartments, where peptide loading is catalyzed by HLA-DM. Disruption of HLA-DRA prevents surface HLA-DR expression, eliminating TCR engagement and CD4+ T cell activation. Downstream effects include impaired IL-2 and IFN-gamma release, compromising adaptive immunity.
In A-549 cells, knockout of HLA-DRA ablates inducible MHC class II expression, offering a clean system to study the role of non-professional antigen presentation in lung adenocarcinoma. This model is instrumental for examining how tumors evade CD4+ T cell responses by downregulating MHC class II, particularly under IFNG exposure. It allows dissection of the CIITA regulatory network and its cross-talk with NSCLC driver mutations. The polyclonal format captures editing heterogeneity, mimicking clonal variation in tumors.
Research applications include co-culture assays with CD4+ T cells to measure T cell activation via IL-2 ELISA, antigen presentation assays using exogenous peptides, and flow cytometric monitoring of HLA-DR surface levels. These cells are suitable for screening small molecules or siRNAs that modulate MHC class II expression and for studying immune evasion mechanisms in NSCLC. RT-qPCR can quantify HLA-DRA mRNA after IFNG stimulation. For further information, please contact Ascent Research.
