# Regulation of RORα Stability through PRMT5-Dependent Symmetric Dimethylation

## TL;DR

This study shows how PRMT5 stabilizes RORα protein through methylation, which may help suppress breast cancer progression.

## Contribution

The study identifies PRMT5-dependent symmetric dimethylation as a novel mechanism regulating RORα stability in breast epithelial cells.

## Key findings

- PRMT5 symmetrically dimethylates the DNA-binding domain of RORα, stabilizing the protein.
- PRMT5 silencing reduces RORα protein levels and increases cell invasion and migration.
- PRMT5 overexpression increases RORα accumulation and suppresses cell invasion.

## Abstract

Retinoid-related orphan receptor alpha (RORα), a member of the orphan nuclear factor family, is considered a potential tumor suppressor. Our previous studies have shown that a loss of RORα is associated with enhanced breast cancer malignancy, cell invasion and proliferation, epithelial–mesenchymal transition, and cancer-associated inflammation. The mechanisms of how RORα expression is regulated in mammary epithelial cells remain incompletely understood. In this study, we revealed a direct interaction between RORα and protein arginine N-methyltransferase 5 (PRMT5), which symmetrically dimethylated the DNA-binding domain of RORα and stabilized the RORα protein. The RORα protein was decreased in PRMT5-silenced mammary epithelial cells, accompanied by enhanced invasion and migration abilities. These findings uncover a novel mechanism for RORα regulation through PRMT5-induced symmetric dimethylation in breast epithelial cells.

Retinoic acid receptor-related orphan receptor alpha (RORα), a candidate tumor suppressor, is prevalently downregulated or lost in malignant breast cancer cells. However, the mechanisms of how RORα expression is regulated in breast epithelial cells remain incompletely understood. Protein arginine N-methyltransferase 5 (PRMT5), a type II methyltransferase catalyzing the symmetric methylation of the amino acid arginine in target proteins, was reported to regulate protein stability. To study whether and how PRMT5 regulates RORα, we examined the direct interaction between RORα and PRMT5 by immunoprecipitation and GST pull-down assays. The results showed that PRMT5 directly bound to RORα, and PRMT5 mainly symmetrically dimethylated the DNA-binding domain (DBD) but not the ligand-binding domain (LBD) of RORα. To investigate whether RORα protein stability is regulated by PRMT5, we transfected HEK293FT cells with RORα and PRMT5-expressing or PRMT5-silencing (shPRMT5) vectors and then examined RORα protein stability by a cycloheximide chase assay. The results showed that PRMT5 increased RORα protein stability, while silencing PRMT5 accelerated RORα protein degradation. In PRMT5-silenced mammary epithelial cells, RORα protein expression was decreased, accompanied by an enhanced epithelial–mesenchymal transition morphology and cell invasion and migration abilities. In PRMT5-overexpressed mammary epithelial cells, RORα protein was accumulated, and cell invasion was suppressed. These findings revealed a novel mechanism by which PRMT5 regulates RORα protein stability.