Reply: Hereditary myopathy with early respiratory failure is caused by mutations in the titin FN3 119 domain

Sir, The letter from Hedberg et al. (2013) is of great interest because it addresses an important question relating to the genetic aetiology of hereditary myopathy with early respiratory failure (HMERF). The original report by Lange et al. (2005) indicated that HMERF was associated with a heterozygous g.296459C4T/ p.R32450W mutation in the kinase domain of titin (TTN) (using Genebank AJ277892 and Uniprot Q8WZ42 as the reference sequences). Since then, no further cases of HMERF caused by kinase domain mutations in TTN have been reported. In contrast, only a year since the report of a mutation in the 119th fibronectin3 (FN3) domain of TTN causing HMERF (Ohlsson et al., 2012; Pfeffer et al., 2012), numerous reports have confirmed an association between the g.274375T4C/p.C30071R FN3 domain mutation and this disease, and seven other mutations have been reported affecting the same domain of titin (Izumi et al., 2013; Palmio et al., 2013; Pfeffer et al., 2013; Toro et al., 2013). Furthermore, our recent study screening 127 patients with myofibrillar myopathy for mutations in TTN revealed seven families with mutations in the 119th FN3 domain, but none with kinase domain mutations (Pfeffer et al., 2013). Ostensibly, the patients from Lange et al.’s (2005) original report were from three separate families, although they all shared the same TTN haplotype (Lange et al., 2005). Therefore, for all intents and purposes, the kinase domain variant has still only been reported in a single HMERF family. Hedberg et al. (2013) now report that patients from the same family described by Lange et al. (2005) not only carry the kinase domain mutation (g.296459C4T/p.R32450W), but also have a g.284762C4T mutation in the 119th fibronectin-3 domain, predicted to cause p.P30091L, which is known to cause HMERF on its own (Palmio et al., 2013; Pfeffer et al., 2013). This finding should be confirmed by the authors of the study by Lange et al. (2005), but assuming that Hedberg et al.’s (2013) report applies to all family members, these data suggest that the g.284762C4T/p.P30091L FN3 domain variant is the true cause of HMERF in the original study of Lange et al. (2005). The segregation of both the FN3 and kinase variants with the disease in these patients is likely because of the shared TTN haplotype between the patients. It should be noted that Lange et al. (2005) presented functional data in support of the kinase variant’s pathogenicity by creating a recombinant TTN kinase construct using previously published methods (Mayans et al., 1998). The catalytic activity of the mutated construct was similar to wild-type, although binding with NBR1 was reduced. This altered NBR1 binding was postulated to be the disease-causing mechanism for this variant. However, the construct included only 324 amino acids from this giant protein, whose inferred complete model includes up to 34 350 amino acids (Uniprot Q8WZ42). We therefore do not know whether this kinase domain variant affects the full-length protein, nor whether the altered NBR1 binding of this particular domain is, in itself, pathogenic. To determine whether kinase domain variants contribute to the pathogenesis, we sequenced the entire kinase domain (according to Uniprot Q8WZ42, including positions p.32115–32496) in 33 HMERF patients from eight families with the g.274375T4C/ p.C30071R mutation in the 119th FN3 domain of TTN (Pfeffer et al., 2012, 2013). Our hypothesis was that amino acid sequence-altering variants (previously reported, or novel) would be more prevalent in patients with HMERF than control subjects, and/or that kinase variants would be present in HMERF patients with more severe phenotype. Our control group consisted of 343 disease controls from 261 pedigrees (70% of whom originate from the UK) who have had their exomes sequenced at our centre. doi:10.1093/brain/awt306 Brain 2014: 137; 1–2 | e271