Inherited peripheral neuropathies are frequent neuromuscular disorders known for their clinical and genetic heterogeneity. In 33 families, we identified 8 mutations in HINT1 (encoding histidine triad nucleotide-binding protein 1) by combining linkage analyses with next-generation sequencing and subsequent cohort screening of affected individuals. Our study provides evidence that loss of functional HINT1 protein results in a distinct phenotype of autosomal recessive axonal neuropathy with neuromyotonia.

Myotonic dystrophy type 1 (DM1) is caused by a highly unstable expansion of CTG repeats in the DMPK gene. Its huge phenotypic variability cannot be explained solely by the repeat number. Recently, variant repeats within the DMPK expansions have emerged as potential disease modifiers. The frequency of variant expanded alleles was estimated in 242 DM1 patients from 174 Serbian families using repeat-primed PCR (RP-PCR). The patterns of variant repeats were determined by direct sequencing of RP-PCR or PCR products. PCR-based southern blot was performed to get insight into the intergenerational mutational dynamics of variant expanded alleles. All patients carrying variant repeats were clinically re-examined. Variant repeats were observed in eight patients from five families (2.9%). They were detected only at the 3' end of DMPK expansions. CCG variant repeats were present in seven patients, either as a part of regular runs of CCGCTG hexamer, individual repeats, or CCG blocks. Analyses of three intergenerational transmissions revealed a considerable stability or likely a contraction of variant expanded alleles. Intriguingly, a decrease in age at onset accompanied these transmissions. Overall, patients were characterized by a milder phenotype and/or some atypical symptoms that could be rather clinically suggestive of myotonic dystrophy type 2. In addition, the first case of de novo CTC variant repeat was observed. Variant repeats might explain a part of the phenotypic variability in a small percent of DM1 patients and likely display a stabilizing effect on the meiotic instability of DMPK expanded alleles.

Global developmental delay (GDD), often accompanied by intellectual disability, seizures and other features is a severe, clinically and genetically highly heterogeneous childhood-onset disorder. In cases where genetic causes have been identified, de novo mutations in neuronally expressed genes are a common scenario. These mutations can be best identified by exome sequencing of parent-offspring trios. De novo mutations in the guanine nucleotide-binding protein, beta 1 (GNB1) gene, encoding the Gβ1 subunit of heterotrimeric G proteins, have recently been identified as a novel genetic cause of GDD. Using exome sequencing, we identified 14 different novel variants (2 splice site, 2 frameshift and 10 missense changes) in GNB1 in 16 pediatric patients. One mutation (R96L) was recurrently found in three ethnically diverse families with an autosomal dominant mode of inheritance. Ten variants occurred de novo in the patients. Missense changes were functionally tested for their pathogenicity by assaying the impact on complex formation with Gγ and resultant mutant Gβγ with Gα. Signaling properties of G protein complexes carrying mutant Gβ1 subunits were further analyzed by their ability to couple to dopamine D1R receptors by real-time bioluminescence resonance energy transfer (BRET) assays. These studies revealed altered functionality of the missense mutations R52G, G64V, A92T, P94S, P96L, A106T and D118G but not for L30F, H91R and K337Q. In conclusion, we demonstrate a pathogenic role of de novo and autosomal dominant mutations in GNB1 as a cause of GDD and provide insights how perturbation in heterotrimeric G protein function contributes to the disease.

CTG expansions in DMPK gene, causing myotonic dystrophy type 1 (DM1), are characterized by pronounced somatic instability. A large proportion of variability of somatic instability is explained by expansion size and patient’s age at sampling, while individual-specific differences are attributed to additional factors. The age at onset is extremely variable in DM1, and inversely correlates with the expansion size and individual-specific differences in somatic instability. Three to five percent of DM1 patients carry repeat interruptions and some appear with later age at onset than expected for corresponding expansion size. Herein, we characterized somatic instability of interrupted DMPK expansions and the effect on age at onset in our previously described patients. Repeat-primed PCR showed stable structures of different types and patterns of repeat interruptions in blood cells over time and buccal cells. Single-molecule small-pool PCR quantification of somatic instability and mathematical modeling showed that interrupted expansions were characterized by lower level of somatic instability accompanied by slower progression over time. Mathematical modeling demonstrated that individual-specific differences in somatic instability had greater influence on age at onset in patients with interrupted expansions. Therefore, repeat interruptions have clinical importance for disease course in DM1 patients due to stabilizing effect on DMPK expansions in somatic cells.

Neuropsychological examinations in myotonic dystrophy (DM) patients show a great variability of results from a condition of intellectual disability to the subtle cognitive impairments. It is unclear if different clusters of neuropsychological deficits appear in different phenotypes of DM, or if there are patients with no cognitive deficit at all. The aim of this study is to assess cognitive impairments among patients with different phenotypes of DM type 1 (DM1) and type 2 (DM2), and to potentially define cognitive clusters in these disorders. Study comprised 101 DM1 and 46 DM2 adult patients who were genetically confirmed. Patients underwent analysis of five cognitive domains (visuospatial, executive, attention, memory and language). Virtually all DM1 patients had cognitive defect with approximately 2-3 cognitive domains affected. On the other hand, one-third of DM2 patients had completely normal neuropsychological findings, and in other two-thirds approximately 1-2 domains were affected. Cluster analysis showed that in both diseases visuospatial and executive dysfunctions seemed to be the main cognitive defects, while memory and language impairments appeared in more severe phenotypes. Our results showed that a single form of DM1 or DM2 may consist of several cognitive clusters. Understanding of cognitive impairments in DM is very important to follow positive and side effects in ongoing and future clinical trials.