Native and recombinant soluble CD23 fragments with IgE suppressive activity

Area of Science:

• Immunology
• Molecular Biology
• Cell Biology

Background:

• Soluble CD23 (sCD23) fragments, including 37,000-25,000 MW forms, are released by CD23-bearing cells and modulate IgE synthesis.
• Previous work showed tunicamycin treatment alters sCD23 activity from IgE potentiation to suppression.

Purpose of the Study:

• To investigate the mechanism behind tunicamycin-induced changes in sCD23 biological activity.
• To characterize the specific CD23 fragments responsible for IgE suppression.

Main Methods:

• Treatment of RPMI-8866 cells and CHO transfectants with tunicamycin.
• Analysis of sCD23 fragment molecular weight (MW) using SDS-PAGE.
• Expression of CD23 mutants lacking N-glycosylation sites or carboxy-terminal amino acids.
• Binding assays with purified native and recombinant 16,000 MW sCD23.
• Measurement of IgE synthesis, including IL-4 stimulated synthesis.

Main Results:

• Tunicamycin treatment of RPMI-8866 cells resulted in the release of 16,000 MW sCD23 fragments.
• These 16,000 MW fragments are generated by truncation of both the N-terminal and carboxy-terminal ends of CD23.
• Experiments with CHO transfectants indicated that carboxy-terminal truncation, not N-glycosylation inhibition, is responsible for 16,000 MW sCD23 formation.
• Highly purified 16,000 MW sCD23 fragments bind to IgE and suppress IgE synthesis, including IL-4-stimulated production.

Conclusions:

• The 16,000 MW sCD23 fragment, generated by carboxy-terminal truncation, is responsible for the suppressive activity on IgE synthesis.
• This finding clarifies the mechanism by which tunicamycin alters CD23 function and identifies a specific sCD23 isoform with potent IgE-suppressive properties.