Human UDP-galactose 4'-epimerase (GALE) is required for cell-surface glycome structure and function

Area of Science:

• Cellular metabolism
• Glycobiology
• Molecular signaling

Background:

• Nucleotide sugars (NSs) are vital metabolites for glycan biosynthesis, crucial for cell physiology.
• Dysregulation of NS levels is linked to various diseases, but mammalian regulation mechanisms are poorly understood.
• UDP-galactose 4 omino-epimerase (GALE) interconverts essential NS pairs, suggesting a key regulatory role.

Purpose of the Study:

• To investigate the role of GALE in regulating NS levels and pathway flux in human cells.
• To determine the impact of GALE deletion on cellular glycosylation and signaling pathways.
• To explore the functional consequences of GALE-mediated NS regulation on cell surface molecules and apoptosis.

Main Methods:

• CRISPR/Cas9 gene editing to create GALE-deficient human cell lines.
• Immunoblotting, flow cytometry, and LC-MS-based profiling for NS, glycolipid, and glycan analysis.
• Assessment of cell surface protein expression and ligand-induced apoptosis.

Main Results:

• GALE deletion led to significant imbalances in nucleotide sugar pools.
• Major alterations in glycolipids and glycoproteins, including integrins and FS-7-associated surface antigen, were observed.
• Decreased levels of sialic acid, galactose, and N-acetylgalactosamine (GalNAc) in glycans were detected.
• GALE-deficient cells showed impaired death receptor signaling and increased apoptosis upon ligand stimulation.

Conclusions:

• GALE plays a critical role in maintaining nucleotide sugar homeostasis and cellular glycosylation.
• GALE-mediated regulation of NS is essential for proper cell surface protein function and death receptor signaling.
• These findings highlight GALE's importance in cell physiology and suggest its potential involvement in diseases associated with NS imbalances, such as galactosemia and metabolic syndrome.