title: Accelerated turnover of MHC class II molecules in nonobese diabetic mice is developmentally and environmentally regulated in vivo and dispensable for autoimmunity.
abstract: The H2-A(g7) (A(g7)) MHC class II (MHCII) allele is required for type 1 diabetes (T1D) in NOD mice. A(g7) not only has a unique peptide-binding profile, it was reported to exhibit biochemical defects, including accelerated protein turnover. Such defects were proposed to impair Ag presentation and, thus, self-tolerance. Here, we report measurements of MHCII protein synthesis and turnover in vivo. NOD mice and BALB/c controls were labeled continuously with heavy water, and splenic B cells and dendritic cells were isolated. MHCII molecules were immunoprecipitated and digested with trypsin. Digests were analyzed by liquid chromatography/mass spectrometry to quantify the fraction of newly synthesized MHCII molecules and, thus, turnover. MHCII turnover was faster in dendritic cells than in B cells, varying slightly between mouse strains. Some A(g7) molecules exhibited accelerated turnover in B cells from young, but not older, prediabetic female NOD mice. This acceleration was not detected in a second NOD colony with a high incidence of T1D. Turnover rates of A(g7) and H2-A(d) were indistinguishable in (NOD × BALB/c) F1 mice. In conclusion, accelerated MHCII turnover may occur in NOD mice, but it reflects environmental and developmental regulation, rather than a structural deficit of the A(g7) allele. Moreover, this phenotype wanes before the onset of overt T1D and is dispensable for the development of autoimmune diabetes. Our observations highlight the importance of in vivo studies in understanding the role of protein turnover in genotype/phenotype relationships and offer a novel approach for addressing this fundamental research challenge.
authors.affiliation: Department of Medicine, University of Cambridge, Addenbrooke's Hospital, Cambridge CB2 0QQ, United Kingdom.
authors.forename: Alessandra
authors.initials: A
authors.lastname: De Riva
authors.forename: Mark C
authors.initials: MC
authors.lastname: Varley
authors.forename: Leslie J
authors.initials: LJ
authors.lastname: Bluck
authors.forename: Anne
authors.initials: A
authors.lastname: Cooke
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Deery
authors.forename: Robert
authors.initials: R
authors.lastname: Busch
mesh_terms: D000818:Animals; D001324:Autoantigens / Q000276:immunology / Q000378:metabolism; D015551:Autoimmunity / Q000502:physiology*; D001402:B-Lymphocytes / Q000276:immunology / Q000378:metabolism; D002853:Chromatography, Liquid; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D003922:Diabetes Mellitus, Type 1 / Q000276:immunology / Q000378:metabolism*; D005434:Flow Cytometry; D000949:Histocompatibility Antigens Class II / Q000276:immunology / Q000378:metabolism*; D047468:Immunoprecipitation; D013058:Mass Spectrometry; D051379:Mice; D008807:Mice, Inbred BALB C; D016688:Mice, Inbred NOD
chemical_list: D001324:Autoantigens; D000949:Histocompatibility Antigens Class II