Structural requirements for C3d,g/Epstein-Barr virus receptor (CR2/CD21) ligand binding, internalization, and viral infection.
The structure of CR2, the human C3d,g/EBV receptor (CR2/CD21) consists of 15 or 16 60-70 amino acid repeats called short consensus repeats (SCRs) followed by a transmembrane and a 34-amino acid intracytoplasmic domain. Functions of CR2 include binding the human complement component C3d,g when it is covalently attached to targets or cross-linked in the fluid phase. In addition, CR2 binds the Epstein-Barr virus (EBV) and mediates internalization of EBV and subsequent infection of cells. In order to explore functional roles of the repetitive extracytoplasmic SCR structure and the intracytoplasmic domain of CR2, we have created truncated CR2 (rCR2) mutants bearing serial deletions of extracytoplasmic SCRs and also the intracytoplasmic tail. We then stably transfected these rCR2 mutants into two cell lines, murine fibroblast L cells and human erythroleukemic K562 cells. Phenotypic analysis of these expressed mutants revealed that 1) The C3d,g- and EBV-binding sites are found in the two amino-terminal SCRs of CR2, 2) expression of SCRs 3 and 4 is further required for high affinity binding to soluble cross-linked C3d,g, 3) the intracytoplasmic domain of CR2 is not required for binding C3d,g or EBV but is necessary for internalization of cross-linked C3d,g as well as for EBV infection of cells, 4) monoclonal anti-CR2 antibodies with similar activities react with single widely separated epitopes, and 5) no functional roles can yet be clearly assigned to SCRs 5-15, as rCR2 mutants not containing these SCRs show no major differences from wild-type rCR2 in binding or internalizing cross-linked C3d,g or mediating EBV binding and infection.
Howard Hughes Medical Institute Laboratories, Washington University School of Medicine, St. Louis, Missouri 63110.
J C
JC
Carel
B L
BL
Myones
B
B
Frazier
V M
VM
Holers
D000818:Animals; D000911:Antibodies, Monoclonal; D000944:Antigens, Differentiation, B-Lymphocyte / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002460:Cell Line; D003176:Complement C3 / Q000378:metabolism*; D003179:Complement C3b; D015933:Complement C3d / Q000378:metabolism*; D004252:DNA Mutational Analysis; D004705:Endocytosis; D000939:Epitopes; D004854:Herpesvirus 4, Human / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009841:Oligonucleotides; D011951:Receptors, Complement / Q000235:genetics / Q000378:metabolism*; D017464:Receptors, Complement 3d; D011991:Receptors, Virus / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014412:Tumor Virus Infections / Q000503:physiopathology
D000911:Antibodies, Monoclonal; D000944:Antigens, Differentiation, B-Lymphocyte; D003176:Complement C3; D000939:Epitopes; D009841:Oligonucleotides; D011951:Receptors, Complement; D017464:Receptors, Complement 3d; D011991:Receptors, Virus; C034772:complement C3g; D003179:Complement C3b; D015933:Complement C3d