LAUSR

involving the proximal muscles in the lower limbs for Family 1, and early 50s with post-exercise myalgia in the lower limbs for Family 2 (Supplementary Table 1). Histological and ultrastructural analyses of the muscle biopsies displayed tubular aggregates as the main histopathological hallmark in both families (Fig. 1a). Exome sequencing identified the heterozygous CASQ1 missense mutations c.166A>T (N56Y) in exon 1 in Family 1, and c.308G>A (G103D) in exon 2 in Family 2. Both mutations affect highly conserved amino acids (Supp. Figure 1), none was found in the available healthy family members, and none was listed in the public or internal SNP databases. A single CASQ1 missense mutation (D244G) has previously been associated with vacuolar myopathy involving protein aggregates [9]. CASQ1 is primarily expressed in skeletal muscle and encodes calsequestrin, the major Ca2+ storage protein in the sarcoplasmic reticulum. Calsequestrin binds Ca2+ with moderate affinity and high capacity, and forms higher order polymers with increasing Ca2+-binding capacities [4]. Immunohistochemistry on a muscle biopsy from Family 2 revealed strong signals for calsequestrin, STIM1, and RyR1 in aggregated structures most likely corresponding to the tubular aggregates, while ORAI1 was not trapped (Fig. 1b). This conforms to the observations made on biopsies from STIM1 and ORAI1 patients and demonstrates that the trapped proteins are primarily of sarcoplasmic reticulum origin [1, 2]. These findings on a single muscle biopsy also suggest that aggregation of STIM1 appears to be a consequence of CASQ1 mutations, providing a pathological link between STIM1and CASQ1-related TAM. In transfected C2C12 myoblasts, WT and both TAM N56Y and G103D mutants formed calsequestrin networks of comparable complexity, while the vacuolar myopathy D244G mutant induced major calsequestrin aggregation (Fig. 1c). Calsequestrin polymerization and depolymerization are dynamic Tubular aggregate myopathy (TAM) is a rare muscle disorder characterized by abnormal accumulations of membrane tubules in muscle fibers, and marked by progressive muscle weakness, cramps, and myalgia [3]. Genetically, TAM has been assigned to mutations in STIM1 [2] and ORAI1 [7], both encoding key regulators of Ca2+ homeostasis. Through exome sequencing of molecularly undiagnosed TAM cases, we now identified CASQ1 as the third TAM gene, and we support our findings by clinical, histological, genetic, and functional data. Family 1 has an ancestral history of a muscle phenotype segregating as a dominant disease, and a partial clinical and histological description was reported earlier [8]. Patient 103901 from Family 2 is a singleton. Birth, early childhood, and motor milestones were normal for all affected members from both families. Disease onset was between early 20s and mid-40s with a slowly progressive muscle weakness mainly