Expression and characterization of recombinant soluble peptide: I-A complexes associated with murine experimental autoimmune diseases.
Structural and functional studies of murine MHC class II I-A molecules have been limited by the low yield and instability of soluble, recombinant heterodimers. In the murine autoimmune diseases experimental autoimmune encephalomyelitis and collagen-induced arthritis, MHC class II molecules I-Au and I-Aq present peptides derived from myelin basic protein and type II collagen, respectively, to autoreactive T cells. To date, systems for the expression of these two I-A molecules in soluble form for use in structure-function relationship studies have not been reported. In the present study, we have expressed functional I-Au and I-Aq molecules using a baculovirus insect cell system. The chain pairing and stability of the molecules were increased by covalently linking the antigenic peptides to beta-chains and adding carboxyl-terminal leucine zippers. Peptide:I-Aq complex quantitatively formed an SDS-stable dimer, whereas peptide:I-Au formed undetectable amounts. However, the two complexes did not show any significant difference in their response to thermal denaturation as assessed by circular dichroism analyses. The autoantigen peptide:I-A complexes were highly active in stimulating cognate T cells to secrete IL-2 and inducing Ag-specific apoptosis of the T cells. Interestingly, the T cells were stimulated by these soluble molecules in the apparent absence of experimentally induced cross-linking of TCRs, indicating that they may have therapeutic potential in autoimmune disease models.
Radu
C G
CG
Department of Microbiology and Cancer Immunobiology Center, University of Texas Southwestern Medical Center, Dallas 75235, USA.
Ober
B T
BT
Colantonio
L
L
Qadri
A
A
Ward
E S
ES
D000818:Animals; D016367:Baculoviridae; D002460:Cell Line; D002942:Circular Dichroism; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism*; D003922:Diabetes Mellitus, Type 1 / Q000378:metabolism*; D019281:Dimerization; D000949:Histocompatibility Antigens Class II / Q000235:genetics / Q000378:metabolism*; D046911:Macromolecular Substances; D051379:Mice; D011994:Recombinant Proteins / Q000378:metabolism; D012967:Sodium Dodecyl Sulfate; D012995:Solubility; D018411:Spodoptera; D013329:Structure-Activity Relationship
D000949:Histocompatibility Antigens Class II; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate