DMPK

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Myotonin-protein kinase (EC 2.7.11.1) (MT-PK) (DM-kinase) (DMK) (DM1 protein kinase) (DMPK) (Myotonic dystrophy protein kinase) [DM1PK] [MDPK]

Publications[править]

Expanded CUG Repeats Trigger Disease Phenotype and Expression Changes through the RNAi Machinery in C. elegans.

Myotonic dystrophy type 1 is an autosomal-dominant inherited disorder caused by the expansion of CTG repeats in the 3' untranslated region of the DMPK gene. The RNAs bearing these expanded repeats have a range of toxic effects. Here we provide evidence from a Caenorhabditis elegans myotonic dystrophy type 1 model that the RNA interference (RNAi) machinery plays a key role in causing RNA toxicity and disease phenotypes. We show that the expanded repeats systematically affect a range of endogenous genes bearing short non-pathogenic repeats and that this mechanism is dependent on the small RNA pathway. Conversely, by perturbating the RNA interference machinery, we reversed the RNA toxicity effect and reduced the disease pathogenesis. Our results unveil a role for RNA repeats as templates (based on sequence homology) for moderate but constant gene silencing. Such a silencing effect affects the cell steady state over time, with diverse impacts depending on tissue, developmental stage, and the type of repeat. Importantly, such a mechanism may be common among repeats and similar in human cells with different expanded repeat diseases.

MeSH Terms

  • 3' Untranslated Regions
  • Aging
  • Animals
  • Animals, Genetically Modified
  • Caenorhabditis elegans
  • Caenorhabditis elegans Proteins
  • Disease Models, Animal
  • Genes, Reporter
  • Green Fluorescent Proteins
  • HSP70 Heat-Shock Proteins
  • Heat-Shock Proteins
  • Hot Temperature
  • Humans
  • Myotonic Dystrophy
  • Protein Binding
  • RNA Interference
  • RNA, Double-Stranded
  • RNA, Helminth
  • Trinucleotide Repeats

Keywords

  • C. elegans
  • RNA interference
  • RNA toxicity
  • myotonic dystrophy
  • trinucleotide repeat disorders


Expanded CTG repeats trigger miRNA alterations in Drosophila that are conserved in myotonic dystrophy type 1 patients.

Myotonic dystrophy type 1 (DM1) is caused by the expansion of CTG repeats in the 3' untranslated region of the DMPK gene. Several missplicing events and transcriptional alterations have been described in DM1 patients. A large number of these defects have been reproduced in animal models expressing CTG repeats alone. Recent studies have also reported miRNA dysregulation in DM1 patients. In this work, a Drosophila model was used to investigate miRNA transcriptome alterations in the muscle, specifically triggered by CTG expansions. Twenty miRNAs were differentially expressed in CTG-expressing flies. Of these, 19 were down-regulated, whereas 1 was up-regulated. This trend was confirmed for those miRNAs conserved between Drosophila and humans (miR-1, miR-7 and miR-10) in muscle biopsies from DM1 patients. Consistently, at least seven target transcripts of these miRNAs were up-regulated in DM1 skeletal muscles. The mechanisms involved in dysregulation of miR-7 included a reduction of its primary precursor both in CTG-expressing flies and in DM1 patients. Additionally, a regulatory role for Muscleblind (Mbl) was also suggested for miR-1 and miR-7, as these miRNAs were down-regulated in flies where Mbl had been silenced. Finally, the physiological relevance of miRNA dysregulation was demonstrated for miR-10, since over-expression of this miRNA in Drosophila extended the lifespan of CTG-expressing flies. Taken together, our results contribute to our understanding of the origin and the role of miRNA alterations in DM1.

MeSH Terms

  • Animals
  • Base Sequence
  • Cells, Cultured
  • Down-Regulation
  • Drosophila Proteins
  • Drosophila melanogaster
  • Female
  • Gene Expression
  • Gene Expression Regulation
  • Humans
  • Life Expectancy
  • Male
  • MicroRNAs
  • Muscle, Skeletal
  • Myotonic Dystrophy
  • Nuclear Proteins
  • Oligonucleotide Array Sequence Analysis
  • Transcriptome
  • Trinucleotide Repeat Expansion


DM1 CTG expansions affect insulin receptor isoforms expression in various tissues of transgenic mice.

Myotonic dystrophy (DM1) is a dominant autosomal multisystemic disorder caused by the expansion of an unstable CTG trinucleotide repeat in the 3' untranslated region of the DMPK gene. Nuclear accumulation of the enlarged CUG-containing DMPK transcripts has a deleterious effect on the regulation of alternative splicing of some RNAs and has a central role in causing the symptoms of DM1. In particular, Insulin Receptor (IR) mRNA splicing defects have been observed in the muscle of DM1 patients. In this study, we have investigated IR splicing in insulin-responsive tissues (i.e. skeletal muscles, adipose tissue, liver) and pancreas and we have studied glucose metabolism in mice carrying the human genomic DM1 region with expanded (>350 CTG) or normal (20 CTG) repeats and in wild-type mice. Mice carrying DM1 expansions displayed a tissue- and age-dependent abnormal regulation of IR mRNA splicing in all the tissues that we investigated. Furthermore, these mice showed a basal hyperglycemia and glucose intolerance which disappeared with age. Our findings show that deregulation of IR splicing due to the DM1 mutation can occur in different mouse tissues, suggesting that CTG repeat expansions might also result in IR misplicing not only in muscles but also in other tissues in DM1 patients.

MeSH Terms

  • Aging
  • Alternative Splicing
  • Animals
  • Gene Expression Profiling
  • Glucose
  • Glucose Tolerance Test
  • Humans
  • Hypothalamus
  • Insulin
  • Insulin Secretion
  • Mice
  • Mice, Transgenic
  • Mutant Proteins
  • Myotonic Dystrophy
  • Myotonin-Protein Kinase
  • Organ Specificity
  • Pancreas
  • Protein Isoforms
  • Protein-Serine-Threonine Kinases
  • RNA, Messenger
  • Receptor, Insulin
  • Transgenes
  • Trinucleotide Repeat Expansion


Fen1 does not control somatic hypermutability of the (CTG)(n)*(CAG)(n) repeat in a knock-in mouse model for DM1.

The mechanism of trinucleotide repeat expansion, an important cause of neuromuscular and neurodegenerative diseases, is poorly understood. We report here on the study of the role of flap endonuclease 1 (Fen1), a structure-specific nuclease with both 5' flap endonuclease and 5'-3' exonuclease activity, in the somatic hypermutability of the (CTG)(n)*(CAG)(n) repeat of the DMPK gene in a mouse model for myotonic dystrophy type 1 (DM1). By intercrossing mice with Fen1 deficiency with transgenics with a DM1 (CTG)(n)*(CAG)(n) repeat (where 104n110), we demonstrate that Fen1 is not essential for faithful maintenance of this repeat in early embryonic cleavage divisions until the blastocyst stage. Additionally, we found that the frequency of somatic DM1 (CTG)(n)*(CAG)(n) repeat instability was essentially unaltered in mice with Fen1 haploinsufficiency up to 1.5 years of age. Based on these findings, we propose that Fen1, despite its role in DNA repair and replication, is not primarily involved in maintaining stability at the DM1 locus.

MeSH Terms

  • Aging
  • Animals
  • Blastocyst
  • DNA Repair
  • DNA Replication
  • Disease Models, Animal
  • Flap Endonucleases
  • Humans
  • Mice
  • Mice, Transgenic
  • Myotonic Dystrophy
  • Quantitative Trait Loci
  • Trinucleotide Repeat Expansion


Transgenic overexpression of human DMPK accumulates into hypertrophic cardiomyopathy, myotonic myopathy and hypotension traits of myotonic dystrophy.

Abnormal expression of human myotonic dystrophy protein kinase (hDMPK) gene products has been implicated in myotonic dystrophy type 1 (DM1), yet the impact of distress accumulation produced by persistent overexpression of this poorly understood member of the Rho kinase-related protein kinase gene-family remains unknown. Here, in the aged transgenic murine line carrying approximately 25 extra copies of a complete hDMPK gene with all exons and an intact promoter region (Tg26-hDMPK), overexpression of mRNA and protein transgene products in cardiac, skeletal and smooth muscles resulted in deficient exercise endurance, an integrative index of muscle systems underperformance. In contrast to age-matched (11-15 months) wild-type controls, hearts from Tg26-hDMPK developed cardiomyopathic remodeling with myocardial hypertrophy, myocyte disarray and interstitial fibrosis. Hypertrophic cardiomyopathy was associated with a propensity for dysrhythmia and characterized by overt intracellular calcium overload promoting nuclear translocation of transcription factors responsible for maladaptive gene reprogramming. Skeletal muscles in distal limbs of Tg26-hDMPK showed myopathy with myotonic discharges coupled with deficit in sarcolemmal chloride channels, required regulators of hyperexcitability. Fiber degeneration in Tg26-hDMPK resulted in sarcomeric disorganization, centralization of nuclei and tubular aggregation. Moreover, the reduced blood pressure in Tg26-hDMPK indicated deficient arterial smooth muscle tone. Thus, the cumulative stress induced by permanent overexpression of hDMPK gene products translates into an increased risk for workload intolerance, hypertrophic cardiomyopathy with dysrhythmia, myotonic myopathy and hypotension, all distinctive muscle traits of DM1. Proper expression of hDMPK is, therefore, mandatory in supporting the integral balance among cytoarchitectural infrastructure, ion-homeostasis and viability control in various muscle cell types.

MeSH Terms

  • Aging
  • Animals
  • Cardiomyopathy, Hypertrophic
  • Echocardiography
  • Exercise Test
  • Gene Dosage
  • Gene Expression
  • Humans
  • Hypotension
  • Mice
  • Mice, Transgenic
  • Muscle, Skeletal
  • Muscle, Smooth, Vascular
  • Myocardium
  • Myotonic Dystrophy
  • Myotonin-Protein Kinase
  • Protein-Serine-Threonine Kinases
  • Transgenes


The pleiotropic expression of the myotonic dystrophy protein kinase gene illustrates the complex relationships between genetic, biological and clinical covariates of male aging.

Aging is a complex process modulated by multiple interactions between environmental and genetic factors. Myotonic dystrophy (DM1) is an autosomal dominant disorder caused by an unstable (CTG)n repeat expansion in the DM1 protein kinase (DMPK) gene. The affected male patients' life expectancy at birth (53.2 years) is more than two decades below that observed in most occidental populations. The DMPK gene expression is pleiotropic and includes the premature expression of several age-related signs, symptoms and metabolic disturbances including hormonal dysfunctions, progressive decrease in muscular mass, presenile cataracts, alopecia, reduced alertness, insulin resistance, dyslipidemia, erectile dysfunction and hypogonadism. The aim of this study was to analyze the relationship between aging covariates and the severity of DM1 expression in 136 DM1 male subjects. DM1 clinical expression was assessed on a validated neuromuscular disability rating scale and was correlated with plasma total testosterone (rs = -0.31, p < 0.001), luteinizing hormone (LH) (rs = 0.52, p < 0.001) and follicle stimulating hormone (FSH) (rs = 0.54, p < 0.001) levels. Following LH releasing hormone stimulation, FSH and LH concentrations increased as a function of DM1 severity (p < 0.05). Muscular disability in DM1 was also positively associated with fasting plasma insulin and triglyceride concentrations (p < 0.05). The association of plasma apolipoprotein B and low-density lipoprotein cholesterol levels with DM1 was not linear across their distribution and tended to reflect cell membrane damage progression. These results suggest that DM1, a simple Mendelian trait, can represent a valuable model to illustrate the complex relationships between variables associated with male aging.

MeSH Terms

  • Adolescent
  • Adult
  • Aged
  • Aging
  • Disability Evaluation
  • Gene Expression
  • Humans
  • Hypothalamo-Hypophyseal System
  • Male
  • Middle Aged
  • Myotonic Dystrophy
  • Myotonin-Protein Kinase
  • Pituitary-Adrenal System
  • Protein-Serine-Threonine Kinases
  • Severity of Illness Index


Developmental expression of myotonic dystrophy protein kinase in brain and its relevance to clinical phenotype.

To investigate the pathophysiologic role of myotonic dystrophy protein kinase (DMPK) in the brain in myotonic dystrophy (MD), the developmental characteristics of DMPK immunoreactivity in the central nervous system and its alteration with disease were studied. Eleven patients' brain with MD (5 congenital form, 6 adult form) were examined by immunohistochemistry using a specific antibody against synthetic DMPK peptides, antipeptide DM1, and compared with 30 control brains, including 16 age-matched controls. In controls, DM1-immunoreactive neurons appeared in the early fetal frontal cortex and cerebellar granule cell layer, persisting through 29 weeks of gestation and then disappearing. In contrast, immunoreactive neurons continued to persist in the cerebral cortex and cerebellar granule cell layer of MD patients. When we counted DM1-immunoreactive neurons, the increase over controls was greater in the congenital form of MD than in the adult form, and was greater in the cerebrum than in the cerebellum in both forms of MD. DM1 immunostaining was predominantly nuclear, mirroring Western blotting of subcellular fractions. Differences in DM1 expression related to development and to the two forms of MD may be closely related to the pathogenesis of mental retardation in this disease.

MeSH Terms

  • Adult
  • Aging
  • Blotting, Western
  • Brain
  • Child, Preschool
  • Female
  • Fetus
  • Frontal Lobe
  • Humans
  • Immunohistochemistry
  • Infant
  • Male
  • Middle Aged
  • Myotonic Dystrophy
  • Myotonin-Protein Kinase
  • Phenotype
  • Protein-Serine-Threonine Kinases
  • Reference Values


Skeletal muscle sodium channel gating in mice deficient in myotonic dystrophy protein kinase.

Myotonic dystrophy, a progressive autosomal dominant disorder, is associated with an expansion of a CTG repeat tract located in the 3'-untranslated region of a serine/threonine protein kinase, DMPK. DMPK modulates skeletal muscle Na channels in vitro, and thus we hypothesized that mice deficient in DMPK would have altered muscle Na channel gating. We measured macroscopic and single channel Na currents from cell-attached patches of skeletal myocytes from mice heterozygous (DMPK( /-)) and homozygous (DMPK(-/-)) for DMPK loss. In DMPK(-/-) myocytes, Na current amplitude was reduced because of reduced channel number. Single channel recordings revealed Na channel reopenings, similar to the gating abnormality of human myotonic muscular dystrophy (DM), which resulted in a plateau of Na current. The gating abnormality deteriorated with increasing age. In DMPK( /-) muscle there was reduced Na current amplitude and increased Na channel reopenings identical to those in DMPK(-/-) muscle. Thus, these mouse models of complete and partial DMPK deficiency reproduce the Na channel abnormality of the human disease, providing direct evidence that DMPK deficiency underlies the Na channel abnormality in DM.

MeSH Terms

  • Aging
  • Animals
  • Ion Channel Gating
  • Membrane Potentials
  • Mice
  • Muscle, Skeletal
  • Myotonic Dystrophy
  • Myotonin-Protein Kinase
  • Patch-Clamp Techniques
  • Protein-Serine-Threonine Kinases
  • Sodium Channels


Progressive atrioventricular conduction block in a mouse myotonic dystrophy model.

Myotonic dystrophy is caused by expansion of a CTG trinucleotide repeat on human chromosome 19, and leads to progressive skeletal myopathy and atrioventricular conduction disturbances. A murine model of myotonic dystrophy has been designed by targeted disruption of the myotonic dystrophy protein kinase (DMPK) gene. The DMPK-deficient mice display abnormalities in A-V conduction characteristics, similar to the human cardiac phenotype. The purpose of this study was to determine whether age-related progression of A-V block occurs in a mouse model of DMPK-deficiency. Surface ECGs and intracardiac electrophysiology (EP) studies were performed in 60 immature and 90 adult homozygous (DMPK-/-), heterozygous (DMPK /-), and wild-type (WT) DMPK / control mice. Complete studies were obtained on 141 of 150 mice. The RR, PR, QRS, and QT intervals were measured on ECG. Sinus node recovery time, AV refractory periods, paced AV Wenckebach and 2:1 block cycle lengths, atrial and ventricular effective refractory periods were compared between genotypes and age groups. There were no differences in ECG intervals or EP findings in the young mutant mice, but progressive PR prolongation in older mice. The A-V conduction defects are also sensitive to DMPK gene dosage. Adult DMPK-/- mice develop 1 degrees, 2 degrees and 3 degrees A-V block, whereas DMPK /- mice develop only 1 degrees heart block. These data demonstrate that both age and DMPK dose are important factors regulating cardiac conduction in myotonic dystrophy. This mouse model of DM is remarkably similar to the human phenotype, with age-related progression in atrioventricular conduction defects.

MeSH Terms

  • Aging
  • Animals
  • Disease Models, Animal
  • Electrocardiography
  • Female
  • Gene Dosage
  • Heart Block
  • Male
  • Mice
  • Mice, Inbred Strains
  • Myotonic Dystrophy
  • Myotonin-Protein Kinase
  • Protein-Serine-Threonine Kinases