Inefficient assembly and intracellular accumulation of antibodies with mutations in V(H) CDR2.
We previously described secretion defects in four mutants of the murine anti-phosphocholine Ab, T15. The mutant heavy (H) chains had amino acid replacements in the V(H) complementarity-determining region 2 (HCDR2) and were expressed at normal intracellular levels. Here, the intracellular fate of the secretion-defective mutant heavy chains was investigated. Metabolic labeling demonstrated that the T15 wild-type Ab was secreted within a 4-h chase. In contrast, the mutant H chains accumulated with intracellular t(1/2) values ranging from 10 to 24 h. The mutant H chains were associated with increased levels of the molecular chaperones BiP and GRP94, and remained endoglycosidase H sensitive, suggesting retention in the endoplasmic reticulum. Assembly of the mutant H chains with T15 light (L) chain was arrested at the H2 and H2L intermediate stages of the T15 wild-type pathway (H2 --> H2L --> H2L2). Even though some assembly with L chain occurred, it was not as a secretion-competent H2L2 Ig moiety. The T15 L chains coexpressed with mutant H chains were degraded efficiently except for a minor L chain population with a long t(1/2) that was apparently protected at the H2L stage. To our knowledge, this is the first study demonstrating that intracellular half-lives of Ig H and L chains can be influenced by somatic mutations in HCDR2.
Martin
T M
TM
Department of Molecular Microbiology and Immunology, Oregon Health Sciences University, Portland 97201, USA.
Wiens
G D
GD
Rittenberg
M B
MB
D000818:Animals; D001324:Autoantigens / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006207:Half-Life; D006596:Hexosaminidases / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D007147:Immunoglobulin Light Chains / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D007700:Kinetics; D016350:Leucine Zippers / Q000235:genetics* / Q000276:immunology; D016296:Mutagenesis; D010449:Peptide Mapping
D001324:Autoantigens; D004268:DNA-Binding Proteins; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D006596:Hexosaminidases