Multiple modes of transcriptional regulation by the nuclear hormone receptor RARγ in human squamous cell carcinoma

Vitamin A metabolism and signaling through nuclear retinoic acid receptors (RARs α,β,γ) regulate embryogenesis, immune functions, and cell differentiation in most cell types. RARγ is highly expressed in stratified squamous epithelial cells of the oral cavity and skin. Although data indicate that RARγ agonism is antitumorigenic in oral cavity squamous cell carcinoma (OCSCC), the specific, primary gene targets of RARγ remain poorly characterized. Here, we define RARγ signaling pathways through integrating genome-wide RARγ binding by Cleavage under Targets and Release Using Nuclease (CUT&RUN), chromatin histone marks, and global transcriptomics ± agonists in human OCSCC cells and in human OCSCC cells with deletion of RARG (gene for RARγ) (RARGKO). Notably, transcripts for some genes associated with stratified squamous cell differentiation, including NOTCH1, NOTCH3, and the NOTCH ligands, JAG2 and DLL1, were reduced in RARGKO without added ligand. Loss of RARγ binding also reduced expression of a broad group of genes that regulates cell identity and extracellular matrix communication, as well as the retinaldehyde reductase, DHRS3, a crucial retinol homeostasis regulating enzyme. We also discovered targets that were directly repressed by RARγ and thus showed higher expression in RARGKO cells. We identified RARG, PPARG, and RXRA as direct RARγ gene targets, indicating that RARγ could control transcription of other genes via regulation of RXRα, a transcription factor with multiple dimerization partners, in OCSCC. Taken together, RARγ-mediated transcriptional regulation is multifaceted and context-dependent. The delineation of these key RARγ targets and signaling pathways should allow the development of new therapeutics for OCSCC.