An In Vitro Model of Charcot-Marie-Tooth Disease Type 4B2 Provides Insight Into the Roles of MTMR13 and MTMR2 in Schwann Cell Myelination

Charcot-Marie-Tooth Disorder Type 4B (CMT4B) is a demyelinating peripheral neuropathy caused by mutations in myotubularin-related (MTMR) proteins 2, 13, or 5 (CMT4B1/2/3), which regulate phosphoinositide turnover and endosomal trafficking. Although mouse models of CMT4B2 exist, an in vitro model would make possible pharmacological and reverse genetic experiments needed to clarify the role of MTMR13 in myelination. We have generated such a model using Schwann cell-dorsal root ganglion (SC-DRG) explants from Mtmr13(−/−) mice. Myelin sheaths in mutant cultures contain outfoldings highly reminiscent of those observed in the nerves of Mtmr13(−/−) mice and CMT4B2 patients. Mtmr13(−/−) SC-DRG explants also contain reduced Mtmr2, further supporting a role of Mtmr13 in stabilizing Mtmr2. Elevated PI(3,5)P(2) has been implicated as a cause of myelin outfoldings in Mtmr2(−/−) models. In contrast, the role of elevated PI3P or PI(3,5)P(2) in promoting outfoldings in Mtmr13(−/−) models is unclear. We found that over-expression of MTMR2 in Mtmr13(−/−) SC-DRGs moderately reduced the prevalence of myelin outfoldings. Thus, a manipulation predicted to lower PI3P and PI(3,5)P(2) partially suppressed the phenotype caused by Mtmr13 deficiency. We also explored the relationship between CMT4B2-like myelin outfoldings and kinases that produce PI3P and PI(3,5)P(2) by analyzing nerve pathology in mice lacking both Mtmr13 and one of two specific PI 3-kinases. Intriguingly, the loss of vacuolar protein sorting 34 or PI3K-C2β in Mtmr13(−/−) mice had no impact on the prevalence of myelin outfoldings. In aggregate, our findings suggest that the MTMR13 scaffold protein likely has critical functions other than stabilizing MTMR2 to achieve an adequate level of PI 3-phosphatase activity.