N-glycosylation regulates ADAM8 processing and activation

Area of Science:

• Biochemistry
• Molecular Biology
• Cancer Research

Background:

• A Disintegrin And Metalloproteinase 8 (ADAM8) is a transmembrane protein highly expressed in aggressive breast tumors and metastases.
• ADAM8 plays critical roles in Triple-Negative breast cancers (TNBCs), a subtype lacking estrogen receptor alpha (ERα).
• While ADAM8 is a glycoprotein, the specific role of its N-glycosylation in its function and processing remained largely uncharacterized.

Purpose of the Study:

• To investigate the role of N-glycosylation in the processing, activation, localization, and stability of ADAM8.
• To determine if N-glycosylation patterns differ between ERα-negative and ERα-positive breast cancer cells.
• To identify specific N-glycosylation sites critical for ADAM8 function.

Main Methods:

• Site-directed mutagenesis was employed to alter identified N-glycosylation sites on human ADAM8.
• Analysis of ADAM8 processing, cell surface localization, and stability was performed using various biochemical and cell biology techniques.
• N-glycosylation types (high mannose vs. complex) were characterized at specific sites.

Main Results:

• N-glycosylation is essential for the correct processing and activation of ADAM8 in ERα-negative breast cancer cells.
• ADAM8 dimers were detected on the surface of ERα-negative cells but not ERα-positive cells, correlating with N-glycosylation.
• Mutations at Asn-91 and Asn-612 impaired processing and cell surface exit, while mutation at Asn-436 reduced stability via lysosomal degradation. Asn-67 had minimal impact.

Conclusions:

• N-glycosylation is a critical post-translational modification regulating ADAM8 processing, cell surface localization, stability, and ultimately, its activity.
• Specific N-glycosylation sites dictate distinct aspects of ADAM8 maturation and function.
• Targeting ADAM8 N-glycosylation may offer a therapeutic strategy for ERα-negative breast cancers.