Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.1 (RNase)
 16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Muscle-specific transcriptional regulation of DMD gene expression has been inferred from both the histopathology of the disease and, more recently, from the use of cDNA sequences to detect DMD gene transcripts by Northern blot, RNase protection, in situ hybridization, and polymerase chain reaction (PCR) analyses. Several muscle-specific genes have been shown to be transcriptionally activated early in myogenesis and a number of cis-acting promoter elements required for muscle-specific transcriptional induction have been described. In this report we review our recent progress on studies of the mechanisms underlying myogenic regulation of dystrophin gene expression. Indirect immunofluorescence has been used to demonstrate that dystrophin is present at the muscle cell surface very early in the myogenic program. The cloning and sequencing of the dystrophin gene promoter reveals the presence of pre-defined muscle-specific cis-acting promoter elements. Functional assays provide evidence that these upstream sequences are capable of regulating DMD gene expression in a cell-and developmental stage-specific manner.
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PMID:Myogenic regulation of dystrophin gene expression. 268 23

Duchenne muscular dystrophy (DMD), a sex-linked degenerative disorder of the muscle, is one of the most common lethal genetic diseases in man. It affects about one male in 3,500, with an estimated one-third of cases being caused by new mutations. A less severe disease, Becker's muscular dystrophy (BMD), maps to the same chromosomal locus and is most probably an allelic form of DMD. Both diseases are sometimes associated with various degrees of mental retardation; the molecular basis of these phenotypes is unknown (for review, see ref. 1). The giant DMD gene spans approximately 2,000 kilobases (kb) (0.05% of the human genome) and encodes a 14-kb mRNA. The tissue-specificity of its expression has not been precisely determined. Monaco et al., using Northern blots, reported expression of the gene in human fetal skeletal muscle and small intestine but not in human fetal brain, or in human cultured myoblasts and transformed B and T cells. More recently, expression was detected in mouse skeletal and cardiac muscle, but not in mouse brain. Here we show, using a ribonuclease protection assay, that the DMD gene is developmentally regulated in rat and mouse myogenic cell cultures, and that it is expressed in rat and mouse striated muscle, in mouse smooth muscle and in rat, mouse and rabbit brain. We could not detect transcripts in other non-muscle tissues.
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PMID:Expression of the putative Duchenne muscular dystrophy gene in differentiated myogenic cell cultures and in the brain. 334 Feb 14

Nineteen serum enzymes from patients with Duchenne muscular dystrophy and asthma, and normal subjects were studied. These enzymes include aminopeptidases, cathepsin C, angiotensin-converting enzyme, serine proteinase, sulphatase, phosphatase, esterases and ribonuclease. The enzymatic changes in dystrophic patients were related to two parameters: severity of the disease as judged from symptomatology, and duration of the disease. Most of the enzyme levels tested were increased in milder cases, but they tended to decrease with severity of the disease. On the other hand, there was a group of enzymes showing just opposite tendencies: serine proteinase, cathepsin C and ribonuclease. Even when viewed from the relationship to duration of the disease, the above mentioned grouping of enzymes was generally valid. Most of the enzyme levels, including those routinely applied as clinical parameters, tended to decrease, logarithmically, with an increase in duration of the disease. On the contrary, some others, including serine proteinase, cathepsin C and ribonuclease, tended to increase toward their control levels. Such tendencies were not found in the patients with asthma. The discrepancy between the above two groups of enzymes may have some implications for the process of protein degradation in dystrophic patients.
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PMID:Two different modes of enzymatic changes in serum with progression of Duchenne muscular dystrophy. 685 Nov 59

Apoptosis was detected in different muscular diseases, including severe dystrophin deficiency, but apoptotic mechanisms are not completely described in adult skeletal muscle. Studying patients affected by Duchenne muscular dystrophy (DMD) and by facio-scapulo-humeral dystrophy (FSHD) we showed an increase of apoptotic myonuclei, bax, and bcl-2-positive myofibers. Positive correlation was detected between apoptotic nuclei and bax expression (p < 0.01). Expression of caspases was analyzed by RNase protection. Caspase transcript was not detected in normal skeletal muscles. DMD muscles expressed caspase 8, 3, 5, 2, 7 and Granzyme B mRNAs. Low levels of caspase 6, 3, and Granzyme B transcripts were detected in FSHD patients. Tissue levels of caspase 3 protein significantly correlated with apoptotic myonuclei (p < 0.05) and with bax expression (p < 0.01). In all DMD cases the activity of caspase 3 was increased, while the FSHD samples were heterogeneous. These data indicate that human skeletal muscle fibers. during the dystrophic process, modulate the expression of caspases and that caspase 3 is involved in myofiber cell death. opening new perspective in the pharmacological treatments of muscular dystrophies, such as the use of caspase inhibitors.
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PMID:Caspase 3 expression correlates with skeletal muscle apoptosis in Duchenne and facioscapulo human muscular dystrophy. A potential target for pharmacological treatment? 1124 14

The development of trans-splicing vectors opens the door for delivering a large therapeutic gene with adeno-associated viral vectors (AAV). One potential application is to deliver the 6 kb mini-dystrophin gene for Duchenne muscular dystrophy (DMD) gene therapy. However, early attempts have been very disappointing because of low transduction efficiency. We have recently identified mRNA accumulation as a critical barrier for the trans-splicing AAV vectors. This barrier can be overcome by rational selection of the gene splitting site. Here we outline a detailed RNase protection assay-based strategy to determine the optimal gene splitting site for the mini-dystrophin gene. We also provide methods to evaluate transduction efficiency of the mini-dystrophin trans-splicing vectors in mdx mouse, a model for DMD.
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PMID:Design of trans-splicing adeno-associated viral vectors for Duchenne muscular dystrophy gene therapy. 1867 29

The mechanisms by which huge human introns are spliced out precisely are poorly understood. We analyzed large intron 7 (110199 nucleotides) generated from the human dystrophin (DMD) pre-mRNA by RT-PCR. We identified branching between the authentic 5' splice site and the branch point; however, the sequences far from the branch site were not detectable. This RT-PCR product was resistant to exoribonuclease (RNase R) digestion, suggesting that the detected lariat intron has a closed loop structure but contains gaps in its sequence. Transient and concomitant generation of at least two branched fragments from nested introns within large intron 7 suggests internal nested splicing events before the ultimate splicing at the authentic 5' and 3' splice sites. Nested splicing events, which bring the authentic 5' and 3' splice sites into close proximity, could be one of the splicing mechanisms for the extremely large introns.
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PMID:Nested introns in an intron: evidence of multi-step splicing in a large intron of the human dystrophin pre-mRNA. 2339 99

We investigated the molecular mechanisms for in-frame skipping of DMD exon 39 caused by the nonsense c.5480T>A mutation in a patient with Becker muscular dystrophy. RNase-assisted pull down assay coupled with mass spectrometry revealed that the mutant RNA probe specifically recruits hnRNPA1, hnRNPA2/B1 and DAZAP1. Functional studies in a human myoblast cell line transfected with DMD minigenes confirmed the splicing inhibitory activity of hnRNPA1 and hnRNPA2/B1, and showed that DAZAP1, also known to activate splicing, acts negatively in the context of the mutated exon 39. Furthermore, we uncovered that recognition of endogenous DMD exon 39 in muscle cells is promoted by FUSE binding protein 1 (FUBP1), a multifunctional DNA- and RNA-binding protein whose role in splicing is largely unknown. By serial deletion and mutagenesis studies in minigenes, we delineated a functional intronic splicing enhancer (ISE) in intron 38. FUBP1 recruitment to the RNA sequence containing the ISE was established by RNA pull down and RNA EMSA, and further confirmed by RNA-ChIP on endogenous DMD pre-mRNA. This study provides new insights about the splicing regulation of DMD exon 39, highlighting the emerging role of FUBP1 in splicing and describing the first ISE for constitutive exon inclusion in the mature DMD transcript.
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PMID:FUBP1: a new protagonist in splicing regulation of the DMD gene. 2566 18

Mutation-induced exon skipping in the DMD gene can modulate the severity of the phenotype in patients with Duchenne or Becker Muscular Dystrophy. These alternative splicing events are most likely the result of changes in recruitment of splicing factors at cis-acting elements in the mutated DMD pre-mRNA. The identification of proteins involved can be achieved by an affinity purification procedure. Here, we provide a detailed protocol for the in vitro RNA binding assay that we routinely apply to explore molecular mechanisms underlying splicing defects in the DMD gene. In vitro transcribed RNA probes containing either the wild type or mutated sequence are oxidized and bound to adipic acid dihydrazide-agarose beads. Incubation with a nuclear extract allows the binding of nuclear proteins to the RNA probes. The unbound proteins are washed off and then the specifically RNA-bound proteins are released from the beads by an RNase treatment. After separation by SDS-PAGE, proteins that display differential binding affinities for the wild type and mutant RNA probes are identified by mass spectrometry.
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PMID:Identification of Splicing Factors Involved in DMD Exon Skipping Events Using an In Vitro RNA Binding Assay. 2906 62