Multiple dimeric forms of human CD69 result from differential addition of N-glycans to typical (Asn-X-Ser/Thr) and atypical (Asn-X-cys) glycosylation motifs.
CD69 is expressed on the surface of all hematopoietically derived leukocytes and is suggested to function as a multipurpose cell-surface trigger molecule important in the development and activation of many different cell types. Human CD69 contains only a single consensus sequence for N-linked oligosaccharide addition within its extracellular domain (Asn-Val-Thr), yet exists as two distinct glycoforms that are assembled together into disulfide-linked homodimers and heterodimers. The molecular basis for human CD69 heterogeneity has remained elusive. In the current report we show that human CD69 glycoforms are generated before the egress of CD69 proteins from the endoplasmic reticulum to the Golgi and are synthesized under conditions where Golgi processing is inhibited, effectively ruling out the possibility that CD69 heterogeneity results from the differential processing of a single glycosylation site in the Golgi complex. Importantly, these data demonstrate that contrary to current belief, not one but two sites for N-glycan addition exist within the human CD69 extracellular domain and identify the second, "cryptic" CD69 N-glycan attachment site as the atypical Cys-containing glycosylation motif, Asn-Ala-Cys. The results in this study provide a molecular basis for human CD69 heterogeneity and show that multiple dimeric forms of human CD69 result from the variable addition of N-glycans to atypical and typical glycosylation motifs within the CD69 extracellular domain.
Vance
B A
BA
Experimental Immunology Branch, NCI, National Institutes of Health, Bethesda, Maryland 20892-1360, USA.
Wu
W
W
Ribaudo
R K
RK
Segal
D M
DM
Kearse
K P
KP
D015703:Antigens, CD / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001216:Asparagine / Q000378:metabolism*; D002474:Cell-Free System; D016384:Consensus Sequence / Q000235:genetics; D004252:DNA Mutational Analysis; D019281:Dimerization; D004220:Disulfides / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D037181:Lectins, C-Type; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011134:Polysaccharides / Q000378:metabolism; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D013601:T-Lymphocytes / Q000378:metabolism; D014158:Transcription, Genetic; D000099192:CD69 Antigens
D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D001216:Asparagine; D004220:Disulfides; D006023:Glycoproteins; D037181:Lectins, C-Type; D011134:Polysaccharides; D000099192:CD69 Antigens