Mutations in the mitofusin 2 (MFN2) gene, which encodes a mitochondrial GTPase mitofusin protein, have recently been reported to cause both Charcot-Marie-Tooth 2A (CMT2A) and hereditary motor and sensory neuropathy VI (HMSN VI). It is well known that HMSN VI is an axonal CMT neuropathy with optic atrophy. However, the differences between CMT2A and HMSN VI with MFN2 mutations remained to be clarified. Therefore, we studied the phenotypic characteristics of CMT patients with MFN2 mutations. Mutations in MFN2 were screened in 62 unrelated axonal CMT neuropathy families. We calculated CMT neuropathy scores (CMTNSs) and functional disability scales (FDSs) to quantify disease severity. Twenty-one patients with the MFN2 mutations were studied by brain MRI. Ten pathogenic mutations were identified in 26 patients from 15 families (24.2%). Six of these mutations had not been reported, and de novo mutations were observed in five families (33.3%). The electrophysiological patterns of affected individuals with the MFN2 mutations were typical of axonal CMT; however, the clinical and electrophysiological characteristics were markedly different in early (<10 years) and late disease-onset (> or =10 years) groups. All patients with an early onset had severe CMTNS (> or =21) and FDS (6 or 7), whereas most patients with late onset had mild CMTNS (< or =10) and FDS (< or =3). We identified two HMSN VI families with the R364W mutation in the early onset group; however, two other families with the same mutation did not have optic atrophy. In addition, two early onset families with R94W mutations, previously reported for HMSN VI, did not have visual impairment. Interestingly, eight patients had periventricular and subcortical hyperintense lesions by brain MRI. In the late-onset group, three patients had sensorineural hearing loss and two had bilateral extensor plantar responses. We found that MFN2 mutations are the major cause of axonal CMT neuropathy, and that they are associated with variable CNS involvements. Phenotypes were significantly different in the early and late disease-onset groups. Our findings suggest that HMSN VI might be a variant of the early onset severe CMT2A phenotype.

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Parkinson's disease (PD) is a difficult disease to diagnose although it is the second most common neurodegenerative disease. Recent studies show that exosome isolated from urine contains LRRK2 or DJ-1, proteins whose mutations cause PD. To investigate a potential use for urine exosomes as a tool for PD diagnosis, we compared levels of LRRK2, α-synuclein, and DJ-1 in urine exosomes isolated from Korean PD patients and non-PD controls. LRRK2 and DJ-1, but not α-synuclein, were detected in the urine exosome samples, as reported previously. We initially could not detect any significant difference in these protein levels between the patient and the control groups. However, when age, disease duration, L-dopa daily dose, and gender were considered as analytical parameters, LRRK2 and DJ-1 protein levels showed clear gender-dependent differences. In addition, DJ-1 level was significantly higher (1.7-fold) in male patients with PD than that in male non-PD controls and increased in an age-dependent manner in male patients with PD. Our observation might provide a clue to lead to a novel biomarker for PD diagnosis, at least in males.

Leucine-rich repeat kinase 2 (LRRK2) has been identified as a causative gene for Parkinson's disease (PD). LRRK2 contains a kinase and a GTPase domain, both of which provide critical intracellular signal-transduction functions. We showed previously that Rab5b, a small GTPase protein that regulates the motility and fusion of early endosomes, interacts with LRRK2 and co-regulates synaptic vesicle endocytosis. Using recombinant proteins, we show here that LRRK2 phosphorylates Rab5b at its Thr6 residue in in vitro kinase assays with mass spectrophotometry analysis. Phosphorylation of Rab5b by LRRK2 on the threonine residue was confirmed by western analysis using cells stably expressing LRRK2 G2019S. The phosphomimetic T6D mutant exhibited stronger GTPase activity than that of the wild-type Rab5b. In addition, phosphorylation of Rab5b by LRRK2 also exhibited GTPase activity stronger than that of the unphosphorylated Rab5b protein. Two assays testing Rab5's activity, neurite outgrowth analysis and epidermal growth factor receptor degradation assays, showed that Rab5b T6D exhibited phenotypes that were expected to be observed in the inactive Rab5b, including longer neurite length and less degradation of EGFR. These results suggest that LRRK2 kinase activity functions as a Rab5b GTPase activating protein and thus, negatively regulates Rab5b signalling.