Identification of Siglec Ligands Using a Proximity Labeling Method.
J Proteome Res
; 16(10): 3929-3941, 2017 10 06.
Article
in En
| MEDLINE
| ID: mdl-28899088
ABSTRACT
Siglecs are a family of receptor-type glycan recognition proteins (lectins) involved in self-nonself discrimination by the immune system. Identification of Siglec ligands is necessary to understand how Siglec-ligand interaction translates into biological outcomes. However, this is challenging because the interaction is weak. To facilitate identification of Siglec ligands, we adopted a proximity labeling method based on the tyramide radicalization principle. Cells that express Siglec ligands were labeled with Siglec-peroxidase complexes and incubated with biotin tyramide and hydrogen peroxide to generate short-lived tyramide radicals that covalently label the proteins near the Siglec-peroxidase complex. A proof-of-principle experiment using CD22 (Siglec-2) probe identified its known ligands on B cells, including CD22 itself, CD45, and IgM, among others, demonstrating the validity of this method. The specificity of labeling was confirmed by sialidase treatment of target cells and using glycan recognition-deficient mutant CD22 probes. Moreover, possible interactions between biotin-labeled proteins were revealed by literature-based protein-protein interaction network analysis, implying the presence of a molecular cluster comprising CD22 ligands. Further application of this method identified CD44 as a hitherto unknown Siglec-15 ligand on RAW264.7-derived osteoclasts. These results demonstrated the utility of proximity labeling for the identification of Siglec ligands, which may extend to other lectins.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
B-Lymphocytes
/
Sialic Acid Binding Immunoglobulin-like Lectins
/
Immune System
/
Lectins
Type of study:
Diagnostic_studies
Limits:
Animals
/
Humans
Language:
En
Journal:
J Proteome Res
Journal subject:
BIOQUIMICA
Year:
2017
Document type:
Article