Increased Expression of Wild-Type or a Centronuclear Myopathy Mutant of Dynamin 2 in Skeletal Muscle of Adult Mice Leads to Structural Defects and Muscle Weakness

Abstract

Dynamin 2 (DNM2) is a large GTPase implicated in many cellular functions, including cytoskeleton regulation and endocytosis. Although ubiquitously expressed, DNM2 was found mutated in two genetic disorders affecting different tissues: autosomal dominant centronuclear myopathy (ADCNM; skeletal muscle) and peripheral Charcot-Marie-Tooth neuropathy (peripheral nerve). To gain insight into the function of DNM2 in skeletal muscle and the pathological mechanisms leading to ADCNM, we introduced wild-type DNM2 (WT-DNM2) or R465W DNM2 (RW-DNM2), the most common ADCNM mutation, into adult wild-type mouse skeletal muscle by intramuscular adeno-associated virus injections. We detected altered localization of RW-DNM2 in mouse muscle. Several ADCNM features were present in RW-DNM2 mice: fiber atrophy, nuclear mislocalization, and altered mitochondrial staining, with a corresponding reduction in specific maximal muscle force. The sarcomere and triad structures were also altered. We report similar findings in muscle biopsy specimens from an ADCNM patient with the R465W mutation. In addition, expression of wild-type DNM2 induced some muscle defects, albeit to a lesser extent than RW-DNM2, suggesting that the R465W mutation has enhanced activity in vivo. In conclusion, we show the RW-DNM2 mutation acts in a dominant manner to cause ADCNM in adult muscle, and the disease arises from a primary defect in skeletal muscle rather than secondary to peripheral nerve involvement. Therefore, DNM2 plays important roles in the maintenance of adult muscle fibers.

Reducing dynamin 2 expression rescues X-linked centronuclear myopathy

“…Moreover, biochemical analysis suggests several DNM2 CNM mutants increase DNM2 GTPase activity and oligomerization (26,27). Furthermore, overexpression of WT DNM2 by AAV or by miR-133a inhibition resulting in increased DNM2 expression promoted muscle defects and centralization of nuclei in mouse muscle (28,30). We thus hypothesized DNM2-CNM mutations are gain-of-function.…”

“…This was supported in vivo by either knockin or overexpression of the most common CNM-DNM2 patient mutation in mice, which induced CNM-like features at adulthood (28,29), indicating that the disease is not due to haploinsufficiency. Overexpression of WT DNM2 also caused similar CNM-like perturbation to the muscle, albeit to a lesser extent (28,30).…”

“…Overactivation of dynamin-mediated membrane fission either by mutations or increased DNM2 levels may thus alter the proper balance for membrane remodeling. While T-tubule defects are a hallmark of CNM and thus regulation of membrane tubulation at this site a tempting hypothesis for explaining the MTM1/DNM2 pathway, dynamin was never observed on T-tubules in normal adult muscle (28,29). At the physiological level, decreased DNM2 strongly reduced or delayed signs of CNM in Mtm1 -/y mice, including muscle mass, maximal force, and triad and nuclei positioning.…”

“…Western blotting was performed as described previously (28). Primary antibodies were as follows: home-made DNM2-R2680 (1:500), DNM2-R2865 (1:500), MTM1-R2827 (1:500), and GAPDH (1:10,000, MAB374; Chemicon); secondary antibodies were as follows: anti-rabbit HRP or anti-mouse HRP (1:10,000, Jackson ImmunoResearch Laboratories).…”

Dynamin 2 homozygous mutation in humans with a lethal congenital syndrome

“…18 In addition, the perinatal syndrome described in our patients, our in situ Dnm2 hybridization observations and knockdown experiments in zebrafish are consistent with Dnm2 having a role during embryonic development. Although features of the CNM pathology can be reproduced both by knock-in and Adeno-associated virus-mediated exogenous expression approaches for the most prevalent ADCNM-causing DNM2 mutation (R465W), 19,20 Durieux et al 20 reported that the homozygous knock-in mice died shortly after birth. These observations highlight the difference in the severity of the pathology at the heterozygous and homozygous state and are consistent with the differences in the phenotypic spectrum observed for the p.Phe379Val heterozygous and homozygous carriers.…”