Internalization of the lymphocytic surface protein CD22 is controlled by a novel membrane proximal cytoplasmic motif.
CD22 is a key receptor on B-lymphocytes that modulates signaling during antigenic stimulation. We have defined a novel cytoplasmic motif in human CD22 that controls its unusually rapid turnover at the plasma membrane. Chimeric and mutated CD22alpha cDNA vectors were constructed and stably transfected in CD22-negative Jurkat T-lymphocytic cells. Two assays were employed to measure CD22alpha internalization: first, cytoplasmic uptake of radioiodinated anti-CD22 monoclonal antibody; and second, lethal targeting of a toxin, saporin, into cells via CD22 using bispecific F(ab')2 ([anti-CD22 x anti-saporin]) antibody. Results showed that CD22alpha lacking a cytoplasmic tail was not internalized and that replacement of the cytoplasmic tail of CD19 with that of CD22alpha resulted in a chimeric molecule that behaved like CD22alpha and internalized rapidly. Step-wise deletion of the cytoplasmic tail of CD22alpha located the internalization motif to a polar region of 11 residues (QRRWKRTQSQQ) proximal to the plasma membrane, a part of the molecule predicted to form a coil or turn structure. Interestingly, additional CD22 mutants showed that the two glutamine residues sandwiching the serine are critical to internalization but that the serine itself is not.
Lymphoma Research Unit, Tenovus Cancer Laboratory, Southampton General Hospital, Tremona Rd., Southampton SO16 6YD, United Kingdom.
C H
CH
Chan
J
J
Wang
R R
RR
French
M J
MJ
Glennie
D015703:Antigens, CD / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018941:Antigens, CD19 / Q000235:genetics / Q000378:metabolism; D000944:Antigens, Differentiation, B-Lymphocyte / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D015815:Cell Adhesion Molecules; D002462:Cell Membrane / Q000378:metabolism*; D004705:Endocytosis; D006801:Humans; D019169:Jurkat Cells; D037102:Lectins; D008214:Lymphocytes / Q000378:metabolism*; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D051918:Sialic Acid Binding Ig-like Lectin 2; D013601:T-Lymphocytes / Q000378:metabolism
D015703:Antigens, CD; D018941:Antigens, CD19; D000944:Antigens, Differentiation, B-Lymphocyte; C497867:CD22 protein, human; D015815:Cell Adhesion Molecules; D037102:Lectins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D051918:Sialic Acid Binding Ig-like Lectin 2