Membrane protein-focused CRISPR screen identifies ATP2A2 as a druggable transcriptional co-regulator of CCND1 (cyclin D1) in lung adenocarcinoma.

[INTRODUCTION] Lung adenocarcinoma (LUAD) is one of the most commonly seen non-small cell lung cancer (NSCLC). Recent progress has highlighted cyclin-dependent kinase (CDK) inhibitors for treating diverse cancers including NSCLC. However, acquired resistance of CDK inhibitors has prompted the exploration of druggable regulatory or compensatory pathways in cell cycle as alternative or combination therapies of CDK inhibitors.

[AIM] This study aims to discover the upstream regulatory proteins in LUAD using a membrane protein-focused CRISPR screen in A549 cells and dissect their functions.

[MATERIALS AND METHODS] The top hit ATP2A2, an endoplasmic reticulum (ER)-localized protein, was analyzed for its function in LUAD using cell proliferation assay, migration and invasion assays and apoptosis assay. The mechanistic studies of ATP2A2 were conducted using RT-qPCR, Western blotting, RNA-Seq, cell cycle assay, protein immunoprecipitation, mass spectrometry, immunofluorescence microscopy and others.

[KEY FINDINGS] The correlation of ATP2A2 expression with NSCLCs was confirmed with public database and collected clinical samples. ATP2A2 promoted LUAD development by involving in cell cycle regulation. ATP2A2 interacted with another ER protein HACD3 and co-regulated the transcription of CCND1 gene (cyclin D1), which forms complex with CDK4/6. ATP2A2 and HACD3 promoted CCND1 expression by activating NF-κB signaling and the nuclear translocation of p65 protein, which is a known transcription factor of CCND1. Curcumin, an ATP2A2-targeting natural product, could inhibit LUAD both in vitro and in xenografted mouse model.

[SIGNIFICANCE] This study revealed ATP2A2-HACD3-NF-κB as a regulatory axis of cyclin D1 expression and shed light on developing ATP2A2-targeted medications for LUAD treatment.