The skeletal muscle
ryanodine receptor gene (
RYR1; OMIM 180901) on chromosome 19q13.1 encodes the skeletal muscle
calcium release channel. To date, more than 25 missense mutations have been identified in
RYR1 and are associated with
central core disease (CCD; OMIM 117000) and/or the
malignant hyperthermia susceptibility phenotype (MHS1; OMIM 145600). The majority of
RYR1 mutations are clustered in the N-terminal hydrophilic domain of the
protein. Only four mutations have been identified so far in the highly conserved C-terminal region encoding the
luminal/transmembrane domain of the
protein which forms the ion pore. Three of these mutations have been found to segregate with pure or mixed forms of CCD. We have screened the C-terminal domain of the
RYR1 gene for mutations in 50 European patients, diagnosed clinically and/or histologically as having CCD. We have identified five missense mutations (four of them novel) in 13 index patients. The mutations cluster in exons 101 and 102 and replace
amino acids which are conserved in all known vertebrate RYR genes. In order to study the functional effect of these mutations, we have immortalized B-lymphocytes from some of the patients and studied their [Ca(2+)](i) homeostasis. We show that lymphoblasts carrying the newly identified
RYR1 mutations exhibit: (i) a release of
calcium from intracellular stores in the absence of any pharmacological activators of RYR; (ii) significantly smaller
thapsigargin-sensitive intracellular
calcium stores, compared to lymphoblasts from control individuals; and (iii) a normal sensitivity of the
calcium release to the RYR inhibitor
dantrolene. Our data suggest the C-terminal domain of
RYR1 as a hot spot for mutations leading to the CCD phenotype. If the functional alterations of mutated RYR channels observed in lymphoblastoid cells are also present in skeletal muscles this could explain the predominant symptom of CCD, i.e. chronic
muscle weakness. Finally, the study of
calcium homeostasis in lymphoblastoid cells naturally expressing
RYR1 mutations offers a novel non-invasive approach to gain insights into the pathogenesis of MH and CCD.