The HCAR2 Knockout T-47D Polyclonal Cells consist of a CRISPR/Cas9-edited polyclonal knockout population targeting the HCAR2 gene. Supplied as a heterogeneous pool, this product circumvents clonal selection biases and provides a representative loss-of-function model for investigating HCAR2-dependent signaling in a breast cancer setting.
The T-47D cell line originates from a pleural effusion of a human ductal carcinoma and maintains expression of estrogen and progesterone receptors while lacking HER2 amplification. This hormone receptor-positive profile makes it a benchmark model for studying ligand-dependent proliferation, endocrine therapy resistance, and the molecular features of luminal A breast cancer. The cells preserve an epithelial phenotype and are routinely employed in nuclear receptor and cancer metabolism research.
HCAR2 (GPR109A) is a Gi/o-coupled receptor that functions as a sensor for key dietary and endogenous metabolites, including nicotinic acid (niacin), the short-chain fatty acid butyrate, and the ketone body beta-hydroxybutyrate. Upon ligand binding, HCAR2 activates the Gi/o protein family, leading to inhibition of adenylyl cyclase (ADCY), a decrease in intracellular cAMP, and reduced PKA activity. Concurrently, the receptor stimulates ERK1/2 phosphorylation and engages the PI3K/Akt pathway. HCAR2 also interacts with beta-arrestin-2, which modulates receptor desensitization and downstream signaling. Through these mechanisms, HCAR2 exerts anti-lipolytic actions and suppresses inflammatory gene expression, thereby regulating lipid oxidation, immune cell function, and mTOR signaling.
In the context of T-47D breast cancer cells, loss of HCAR2 eliminates the ability to respond to metabolites that normally activate this receptor, offering a unique system to dissect the intersection of metabolite sensing and hormone receptor signaling. Given that HCAR2 activation typically dampens lipolysis and inflammation, its knockout may lead to altered lipid metabolism and a shift in the secretome profile, potentially impacting tumor cell proliferation, migration, and immune evasion. This model is especially valuable for studying how butyrate, a gut microbiota product, influences breast cancer cell behavior via HCAR2, and for exploring potential crosstalk with estrogen-mediated pathways.
These polyclonal knockout cells are suitable for a broad range of functional assays. Researchers can validate HCAR2 disruption via western blotting or RT-qPCR, measure cAMP accumulation following niacin/butyrate stimulation, and assess downstream signaling using phospho-ERK detection. The cells are amenable to cell proliferation, migration, and invasion assays, as well as lipid droplet staining to monitor lipid accumulation. Cytokine profiling by ELISA or transcriptomic analysis after metabolite challenge can reveal HCAR2-dependent inflammatory responses. Co-immunoprecipitation experiments can confirm the loss of Gi/o protein interaction. The model also supports high-throughput screening for GPCR modulators and anti-inflammatory compounds. For additional technical information, please contact Ascent Research.
