Role of the eye muscles in thyroid eye disease: Identification of the principal autoantigens

Thyroid-associated ophthalmopathy (TAO) is a progressive orbital disorder associated with Graves' hyperthyroidism and, less often, Hashimoto's thyroiditis in which autoantibodies react with orbital antigens and lead to exophthalmos and eye muscle inflammation. Eye muscle (EM) membrane proteins initially reported as 55 and 64 kd are the best markers of ophthalmopathy. The "64-kd protein" is now shown to be the flavoprotein subunit of mitochondrial succinate dehydrogenase and to have a correct molecular weight of 67 kd. We have cloned a fragment of a novel eye muscle protein, which we call G2s, and sequenced 1.4 kb of the full length cDNA. G2s does not share any significant homologies with other reported proteins. The 5.9 kb G2s mRNA, that corresponds to a protein of approximately 220 kd, is expressed in EM, other skeletal muscle and thyroid, but not in other tissues tested. We have also cloned and sequenced a 63-kd eye muscle protein identified as the calcium binding protein calsequestrin. Antibodies against calsequestrin were found in 40% of patients with active ophthalmopathy, but in 0% of normal subjects. Finally, we have sequenced a 19 amino acid fragment of a 55-kd porcine eye muscle membrane protein that exactly matched porcine and human sarcalumenin, a 160-kd glycoprotein localized in the lumen of the longitudinal sarcoplasmic reticulum of the skeletal muscle fiber where it binds calcium. A 53-kd glycoprotein fragment of the molecule corresponds to the 55-kd protein. In a preliminary study, serum antibodies against purified sarcalumenin were detected in 40% of patients with active TAO of less than 1 year duration, but in no controls tested. We propose that the primary autoantigen in TAO is G2s, which would also explain the association of ophthalmopathy with thyroid autoimmunity, and that antibodies against the intracellular proteins flavoprotein, calsequestrin, and sarcalumenin are secondary markers of an immune-mediated reaction in eye muscle in patients with thyroid autoimmunity.