A Functional Relationship Between UNC45A and MYO5B Connects Two Rare Diseases With Shared Enteropathy

Area of Science:

• Cell biology
• Molecular genetics
• Gastroenterology

Background:

• Mutations in UNC45A cause osteo-oto-hepato-enteric (O2HE) syndrome, characterized by congenital diarrhea and intrahepatic cholestasis.
• Similar symptoms are observed in microvillus inclusion disease (MVID), linked to mutations in MYO5B, which encodes myosin Vb.
• The functional relationship between UNC45A and myosin Vb in these related enteropathies is unexplored.

Purpose of the Study:

• To investigate the functional link between UNC45A and myosin Vb.
• To determine if UNC45A and myosin Vb interact in intestinal and hepatic cells.
• To elucidate the molecular basis of O2HE syndrome associated with UNC45A mutations.

Main Methods:

• CRISPR-Cas9 gene editing and site-directed mutagenesis in intestinal and hepatocellular cell lines.
• Analysis of protein expression and localization using Western blotting, qPCR, and microscopy.
• Functional assays assessing recycling endosome positioning and microvilli development.

Main Results:

• UNC45A depletion reduced myosin Vb protein levels and impaired myosin Vb-dependent processes (recycling endosome positioning, microvilli development) in intestinal cells.
• Reintroduction of UNC45A restored myosin Vb expression and cellular functions.
• The O2HE-associated UNC45A variant (p.V423D) reduced UNC45A protein stability but did not affect its ability to promote myosin Vb expression or microvilli development.

Conclusions:

• A functional link exists between UNC45A and myosin Vb, connecting O2HE syndrome and MVID at a molecular level.
• The pathogenicity of the O2HE syndrome-associated UNC45A mutation is primarily due to protein instability.
• UNC45A plays a crucial role in maintaining myosin Vb function and intestinal epithelial cell structure.