Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
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Hexokinase coding DM1 and DM2 sequences were obtained from genomic DNA of a Drosophila melanogaster cell line by PCR amplification strategy. Both the sequences were found to encode an enzyme with a molecular weight of 50,000 Da. Amino acid sequence alignment of DM1 and DM2 shows approximately 45% homology with yeast and human hexokinases. The sequences also indicated the presence of conserved amino acid residues and motifs that are present in mammalian hexokinases and are involved in the binding of different substrates. Southern blot analysis suggests that the D. melanogaster genome contain a single copy of DM1 and DM2 sequences. Northern analysis indicates DM1 is expressed as more than one transcript in adult as well as in the D.Mel2 cell line. DM2 is expressed as a single transcript in adult flies. Expression levels for DM1 and DM2 encoded message were found to be similar in different stages of development as seen by RT-PCR. The biotechnological significance of these sequences in metabolic engineering of cells is discussed.
Insect Biochem Mol Biol 2001 Nov 01
PMID:Cloning of two hexokinase isoenzyme sequences from Drosophila melanogaster. 1158 29

In this study we have developed an in vitro cell culture system which displays the majority of the defects previously described for congenital myotonic dystrophy (CDM) muscle in vivo. Human satellite cells were isolated from the quadriceps muscles of three CDM fetuses with different clinical severity. By Southern blot analysis all three cultures were found to have approximately 2300 CTG repeats. This CTG expansion was found to progressively increase in size during the proliferative life span, confirming an instability of this triplet in skeletal muscle cells. The CDM myoblasts and myotubes also showed abnormal retention of mutant RNA in nuclear foci, as well as modifications in their myogenic program. The proliferative capacity of the CDM myoblasts was reduced and a delay in fusion, differentiation and maturation was observed in the CDM cultures compared with unaffected myoblast cultures. The clinical severity and delayed maturation observed in the CDM fetuses were closely reflected by the phenotypic modifications observed in vitro. Since the culture conditions were the same, this suggests that the defects we have described are intrinsic to the program expressed by the myoblasts in the absence of any trophic factors. Altogether, our results demonstrate that satellite cells are defective in CDM and are probably implicated in the delay in maturation and muscle atrophy that has been described previously in CDM fetuses.
Hum Mol Genet 2001 Sep 15
PMID:Defective satellite cells in congenital myotonic dystrophy. 1159 Jan 25

Intraneuronal aggregates of hyperphosphorylated tau proteins, referred to as pathological tau, are found in brain areas of demented patients affected by numerous different neurodegenerative disorders. We previously described a particular biochemical profile of pathological tau proteins in myotonic dystrophy type 1 (DM1). This multisystemic disorder is characterized by an unstable CTG repeat expansion in the 3'-untranslated region of the DM protein kinase gene. In the human central nervous system, tau proteins consist of six isoforms that differ by the presence or absence of the alternatively spliced exons 2, 3 and 10. Here we show that the pattern of tau isoforms aggregated in DM1 brain lesions is characteristic. It consists mainly of the aggregation of the shortest human tau isoform. A disruption in normal tau isoform expression consisting of a reduced expression of tau isoforms containing the exon 2 was observed at both the mRNA and protein levels. Large expanded CTG repeats were detected and showed marked somatic heterogeneity between DM1 cases and in cortical brains regions analysed. Our data suggest a relationship between the CTG repeat expansion and the alteration of tau expression showing that DM1 is a peculiar tauopathy.
Hum Mol Genet 2001 Sep 15
PMID:Dysregulation of human brain microtubule-associated tau mRNA maturation in myotonic dystrophy type 1. 1159 Jan 31

The phenotypes in myotonic dystrophy types 1 and 2 (DM1 and DM2) are similar, suggesting a shared pathophysiologic mechanism. DM1 is caused by expansion of a CTG repeat in the DMPK gene. Pathogenic effects of this mutation are likely to be mediated, at least in part, by the expanded CUG repeat in mutant mRNA. The mutant transcripts are retained in the nucleus in multiple discrete foci. We investigated the possibility that DM2 is also caused by expansion of a CTG repeat or related sequence. Analysis of DNA by repeat expansion detection methods, and RNA by ribonuclease protection, did not show an expanded CTG or CUG repeat in DM2. However, hybridization of muscle sections with fluorescence-labeled CAG-repeat oligonucleotides showed nuclear foci in DM2 similar to those seen in DM1. Nuclear foci were present in all patients with symptomatic DM1 (n = 9) or DM2 (n = 9) but not in any disease controls or healthy subjects (n = 23). The foci were not seen with CUG- or GUC-repeat probes. Foci in DM2 were distinguished from DM1 by lower stability of the probe-target duplex, suggesting that a sequence related to the DM1 CUG expansion accumulates in the DM2 nucleus. Muscleblind proteins, which interact with expanded CUG repeats in vitro, localized to the nuclear foci in both DM1 and DM2. These results support the idea that nuclear accumulation of mutant RNA is pathogenic in DM1, suggest that a similar disease process occurs in DM2, and point to a role for muscleblind in the pathogenesis of both disorders.
Hum Mol Genet 2001 Sep 15
PMID:Muscleblind localizes to nuclear foci of aberrant RNA in myotonic dystrophy types 1 and 2. 1159 Jan 33

Myotonic dystrophy (DM), an autosomal dominant neuromuscular disease, is associated with expansion of a polymorphic (CTG)n repeat in the 3'-untranslated region of the DM protein kinase (DMPK) gene. The repeat expansion results in decreased levels of DMPK mRNA and protein, but the mechanism for this decreased expression is unknown. Loss of a nuclease-hypersensitive site in the region of the repeat expansion has been observed in muscle and skin fibroblasts from DM patients, indicating a change in local chromatin structure. This change in chromatin structure has been proposed as a mechanism whereby the expression of DMPK and neighboring genes, sine oculis homeobox (Drosophila) homolog 5 (SIX5) and dystrophia myotonica-containing WD repeat motif (DMWD), might be affected. We have developed a polymerase chain reaction (PCR)-based method to assay the chromatin sensitivity of the region adjacent to the repeat expansion in somatic cell hybrids carrying either normal or affected DMPK alleles and show that hybrids carrying expanded alleles exhibit decreased sensitivity to PvuII digestion in this region. Semiquantitative multiplex reverse transcriptase PCR (RT/PCR) assays of gene expression from the chromosomes carrying the expanded alleles showed marked reduction of DMPK mRNA, partial inhibition of SIX5 expression from a congenital DM chromosome, and no reduction of DMWD mRNA. Nested RT/PCR analysis of DMPK mRNA from somatic cell hybrids carrying the repeat expansions revealed that most of the DMPK transcripts expressed from the expanded alleles lacked exons 13 and 14, whereas full-length transcripts were expressed predominantly from the normal alleles. These results suggest that the CTG repeat expansion leads to a decrease in DMPK mRNA levels by affecting splicing at the 3' end of the DMPK pre-mRNA transcript.
Mol Genet Metab
PMID:Effect of triplet repeat expansion on chromatin structure and expression of DMPK and neighboring genes, SIX5 and DMWD, in myotonic dystrophy. 1159 25

The autosomal dominant mutation causing myotonic dystrophy (DM1) is a CTG repeat expansion in the 3'-UTR of the DM protein kinase (DMPK) gene. This multisystemic disorder includes myotonia, progressive weakness and wasting of skeletal muscle and extramuscular symptoms such as cataracts, testicular atrophy, endocrine and cognitive dysfunction. The mechanisms underlying its pathogenesis are complex. Recent reports have revealed that DMPK gene haploinsufficiency may account for cardiac conduction defects whereas cataracts may be due to haploinsufficiency of the neighboring gene, the DM-associated homeobox protein (DMAHP or SIX5) gene. Furthermore, mice expressing the CUG expansion in an unrelated mRNA develop myotonia and myopathy, consistent with an RNA gain of function. We demonstrated that transgenic mice carrying the CTG expansion in its human DM1 context (>45 kb) and producing abnormal DMPK mRNA with at least 300 CUG repeats, displayed clinical, histological, molecular and electrophysiological abnormalities in skeletal muscle consistent with those observed in DM1 patients. Like DM1 patients, these transgenic mice show abnormal tau expression in the brain. These results provide further evidence for the RNA trans-dominant effect of the CUG expansion, not only in muscle, but also in brain.
Hum Mol Genet 2001 Nov 01
PMID:Mice transgenic for the human myotonic dystrophy region with expanded CTG repeats display muscular and brain abnormalities. 1172 59

The mechanism of expansion of the (CTG)n repeat in myotonic dystrophy (DM1) patients and the cause of its pathobiological effects are still largely unknown. Most likely, long repeats exert toxicity at the level of nuclear RNA transport or splicing. Here, we analyse cis- and trans-acting parameters that determine repeat behaviour in novel mouse models for DM1. Our mice carry 'humanized' myotonic dystrophy protein kinase (Dmpk) allele(s) with either a (CTG)84 or a (CTG)11 repeat, inserted at the correct position into the endogenous DM locus. Unlike in the human situation, the (CTG)84 repeat in the syntenic mouse environment was relatively stable during intergenerational segregation. However, somatic tissues showed substantial repeat expansions which were progressive upon aging and prominent in kidney, and in stomach and small intestine, where it was cell-type restricted. Other tissues examined showed only marginal size changes. The (CTG)11 allele was completely stable, as anticipated. Introducing the (CTG)84 allele into an Msh3-deficient background completely blocked the somatic repeat instability. In contrast, Msh6 deficiency resulted in a significant increase in the frequency of somatic expansions. Competition of Msh3 and Msh6 for binding to Msh2 in functional complexes with different DNA mismatch-recognition specificity may explain why the somatic (CTG)n expansion rate is differentially affected by ablation of Msh3 and Msh6.
Hum Mol Genet 2002 Jan 15
PMID:Somatic expansion behaviour of the (CTG)n repeat in myotonic dystrophy knock-in mice is differentially affected by Msh3 and Msh6 mismatch-repair proteins. 1180 28

Precise measurement of germline repeat number mutations is important for understanding the molecular etiology of expanded trinucleotide repeat diseases. We used single genome-equivalent PCR of sperm DNA to measure the mutation frequencies in two lines of Dmt transgenic mice containing an expanded CTG.CAG tract on an identical genetic background. Single genome-equivalent PCR indicated that apparent mutational spectra derived in other investigations from PCR of bulk sperm DNA were largely the consequence of PCR stutter and not mutations. Here we show that sperm from 8-week-old Dmt-D mice had a significantly higher mutation frequency (change of >1 repeat) (14.2%) than those of Dmt-E mice of the same age (5.5%), in agreement with pedigree analysis. Furthermore, the mutation frequency in sperm of Dmt-D mice increased significantly with age (28.0% at 17 weeks). The age dependence of the degree of expansion implies that mutations accumulate with time in spermatogenic stem cells. Similar rates of expansion per spermatogenic cycle in man would yield the large expansions observed in human diseases such as myotonic dystrophy type 1. Pedigree data showed a significant age-dependent bias toward repeat contraction in female transmissions and a trend towards expansion with age in male transmissions. Thus, direct single genome-equivalent PCR of the sperm DNA of an individual male appears to predict the distribution of mutant allele sizes that might be inherited by its offspring. In further contrast to a recent report, the sex of the offspring had no detectable effect on the direction of the mutational length change.
Hum Mol Genet 2002 Apr 01
PMID:Age and insertion site dependence of repeat number instability of a human DM1 transgene in individual mouse sperm. 1192 51

Myotonic dystrophy is a complex neuromuscular disorder associated with DNA expansion mutations in two different genes. In DM1 a CTG repeat in the 3'-untranslated region of DMPK is expanded, whereas in DM2 an intronic CCTG expansion occurs in the gene ZNF9. Transcripts containing expanded repeats form foci in the nuclei of DM1 and DM2 cells. Recent work using antibodies has shown that proteins related to Drosophila muscleblind co-localize with repeat foci in DM1 and DM2 cells. We show that rather than there being a single human muscleblind gene producing multiple proteins through alternative splicing, there are in fact three different muscleblind genes, MBNL, MBLL and MBXL, which map to chromosomes 3, 13 and X, respectively, and which show extensive alternative splicing. Two of the genes, MBNL and MBLL, are expressed in many adult tissues whereas MBXL is expressed predominantly in the placenta. Green fluorescent protein-tagged versions of MBNL, MBLL and MBXL co-localize with nuclear foci in DM1 and DM2 cells, suggesting that all three proteins may play a role in DM pathophysiology.
Hum Mol Genet 2002 Apr 01
PMID:Three proteins, MBNL, MBLL and MBXL, co-localize in vivo with nuclear foci of expanded-repeat transcripts in DM1 and DM2 cells. 1192 53

Facioscapulohumeral muscular dystrophy (FSHD) is the third most common inherited neuromuscular disorder after Duchenne muscular dystrophy and myotonic dystrophy. The gene underlying FSHD was mapped to chromosome 4q35 in 1990 and was shown to be closely linked to locus D4F104S1. Although D4F104S1-associated deletions are closely associated with FSHD, the identity and location of the FSHD gene (or genes) still remain elusive, as does the mechanistic basis of the disease. In addition, although approximately 5% of FSHD families fail to exhibit linkage to 4q35, a putative second locus remains unidentified. The search for the FSHD gene has been hampered both by sequence homologies between the 4q35 candidate region and other chromosomal regions and by the presence of many highly repetitive sequences. Molecular diagnosis for FSHD is usually offered with 98% accuracy but because of its complexity, a much more simple test would be preferable. Indeed, the identification of the FSHD gene itself should potentiate major improvements in diagnostic testing.
Expert Rev Mol Diagn 2002 Mar
PMID:Molecular diagnosis of facioscapulohumeral muscular dystrophy. 1196 36


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