The B cell coreceptor CD22 associates with AP50, a clathrin-coated pit adapter protein, via tyrosine-dependent interaction.
The B cell coreceptor CD22 plays an important role in regulating signal transduction via the B cell Ag receptor. Studies have shown that surface expression of CD22 can be modulated in response to binding of ligand (i.e., mAb). Thus, it is possible that alterations in the level of CD22 expression following binding of natural ligand(s) may affect its ability to modulate the Ag receptor signaling threshold at specific points during B cell development and differentiation. Therefore, it is important to delineate the physiologic mechanism by which CD22 expression is controlled. In the current study, yeast two-hybrid analysis was used to demonstrate that CD22 interacts with AP50, the medium chain subunit of the AP-2 complex, via tyrosine-based internalization motifs in its cytoplasmic domain. This interaction was further characterized using yeast two-hybrid analysis revealing that Tyr(843) and surrounding amino acids in the cytoplasmic tail of CD22 comprise the primary binding site for AP50. Subsequent studies using transfectant Jurkat cell lines expressing wild-type or mutant forms of CD22 demonstrated that either Tyr(843) or Tyr(863) is sufficient for mAb-mediated internalization of CD22 and that these motifs are involved in its interaction with the AP-2 complex, as determined by coprecipitation of alpha-adaptin. Finally, experiments were performed demonstrating that treatment of B cells with either intact anti-Ig Ab or F(ab')(2) blocks ligand-mediated internalization of CD22. In conclusion, these studies demonstrate that internalization of CD22 is dependent on its association with the AP-2 complex via tyrosine-based internalization motifs.
John
BinuJoy
B
Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX 77555, USA.
Herrin
Brantley R
BR
Raman
Chander
C
Wang
Yan-ni
YN
Bobbitt
Kevin R
KR
Brody
Brian A
BA
Justement
Louis B
LB
D033962:Adaptor Protein Complex 2 / Q000235:genetics / Q000378:metabolism*; D033983:Adaptor Protein Complex mu Subunits / Q000235:genetics / Q000378:metabolism*; D020816:Amino Acid Motifs / Q000235:genetics; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D015703:Antigens, CD / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000944:Antigens, Differentiation, B-Lymphocyte / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015815:Cell Adhesion Molecules; D002966:Clathrin / Q000378:metabolism* / Q000502:physiology; D003432:Cross-Linking Reagents / Q000378:metabolism; D004705:Endocytosis / Q000235:genetics / Q000276:immunology; D006801:Humans; D019169:Jurkat Cells; D037102:Lectins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D011485:Protein Binding / Q000235:genetics; D011947:Receptors, Antigen, B-Cell / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D051918:Sialic Acid Binding Ig-like Lectin 2; D015398:Signal Transduction / Q000235:genetics / Q000276:immunology; D013154:Spleen / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
D033962:Adaptor Protein Complex 2; D033983:Adaptor Protein Complex mu Subunits; D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D000944:Antigens, Differentiation, B-Lymphocyte; C497867:CD22 protein, human; C497868:Cd22 protein, mouse; D015815:Cell Adhesion Molecules; D002966:Clathrin; D003432:Cross-Linking Reagents; D037102:Lectins; D011947:Receptors, Antigen, B-Cell; D051918:Sialic Acid Binding Ig-like Lectin 2; C438885:adaptor protein complex 2, mu 1 subunit; D014443:Tyrosine