1,747 research outputs found
Structural study of skeletal muscle fibres in healthy and pseudomyotonia affected cattle
No full textCattle congenital pseudomyotonia (PMT), recognized as naturally occurring animal model of human Brody disease, is an inherited recessive autosomal muscular disorder due to missense mutations in ATP2A1 gene, encoding sarco(endo)plasmic reticulum Ca2(+)-ATPase protein, isoform 1 (SERCA1). PMT has been described in the Chianina and Romagnola italian cattle breeds and as a single case in Dutch improved Red and White cross-breed. The genetic defect turned out to be heterogeneous in different cattle breeds, even though clinical symptoms were homogeneous. Skeletal muscles of affected animals are characterized by a selective deficiency of SERCA1 in sarcoplasmic reticulum (SR) membranes. Recently, we provided evidence that in Chianina breed, the ubiquitin proteasome system is responsible for SERCA1 mutant premature disposal, even when the mutation does not affect the catalytic properties of the pump.
Results presented here show that all SERCA1 mutants described until now, although expressed at low level, are correctly targeted to SR membranes. Ultrastructural studies confirm that in pathological muscle fibres, structure, as well as triads, is well preserved.
All together these results suggest that a possible therapeutical approach based on the rescue of the defective protein at SR membranes could be hypothesized. Only fully functionally active missense mutants, whem located at the SR membrane could restore the efficient control of Ca2+ homeostasis and prevent the appearance of the pathological signs. Moreover, these data demonstrate the increasing importance of domestic animals as genetic models of human pathologies
Staley, Roberta
No full textcurrentAcademic Biography
BA (University of Calgary)
Diploma Journalism (Grant MacEwan)
MA Liberal Studies (Simon Fraser University)
Roberta Staley is an author, a magazine editor and writer, and a documentary filmmaker who has reported from such places as Afghanistan, Papua New Guinea, Kenya, El Salvador, Haiti, Colombia, Cambodia, South Africa, Israel, and New Zealand. She currently edits Enterprise magazine, and is a contributor to BC Business, the South China Morning Post Magazine, Ms. Magazine, Trek, the Canadian Chemical News, Corporate Knights, and Sculpture, among others. She is also a columnist for Just for Canadian Doctors/Dentists magazines. Roberta has published her first book, titled Voice of rebellion : how Mozhdah Jamalzadah brought hope to Afghanistan. It is a biography of Afghan-Canadian human rights activist Mozhdah Jamalzadah
Variation of phospholamban in slow-twitch muscle sarcoplasmic reticulum between mammalian species and a link to the substrate specificity of endogenous Ca2+-calmodulin-dependent protein kinase
No full textSystematic immunological and biochemical studies indicate that the level of expression of sarcoplasmic reticulum (SR) Ca(2+)-ATPase regulatory protein phospholamban (PLB) in mammalian slow-twitch fibers varies from zero, in the rat, to significant levels in the rabbit, and even higher in humans. The lack of PLB expression in the rat, at the mRNA level, is shown to be exclusive to slow-twitch skeletal muscle, and not to be shared by the heart, thus suggesting a tissue-specific, in addition to a species-specific regulation of PLB. A comparison of sucrose density-purified SR of rat and rabbit slow-twitch muscle, with regard to protein compositional and phosphorylation properties, demonstrates that the biodiversity is two-fold, i.e. (a) in PLB membrane density; and (b) in the ability of membrane-bound Ca(2+)-calmodulin (CaM)-dependent protein kinase II to phosphorylate both PLB and SERCA2a (slow-twitch isoform of Ca(2+)-ATPase). The basal phosphorylation state of PLB at Thr-17 in isolated SR vesicles from rabbit slow-twitch muscle, colocalization of CaM K II with PLB and SERCA2a at the same membrane domain, and the divergent subcellular distribution of PKA, taken together, seem to argue for a differential heterogeneity in the regulation of Ca(2+) transport between such muscle and heart muscle
Natural mutation in bovine Sarco(endo)plasmatic Reticulum Ca2+ATPase1 (SERCA1): histological and biochemical aspect in the muscle fibers
No full textCongenital Pseudomyotonia (PMT) is an inherited muscular disease affecting bovine species. Clinically it is characterized by an exercise-induced muscle contraction. The aetiology is related to a prolonged high level of cytosolic free Ca2+ ion in muscle fibers. Genetic analysis has provided evidence of mutations in ATP2A1 gene coding for Sarco(endo)plasmic Reticulum Ca2+-ATPase, isoform1 (SERCA1) a membrane protein involved in re-uptake of calcium from cytosol into sarcoplasmic reticulum [1].
The clinical symptoms and genetic correlations make bovine PMT the true counterpart of human Brody’s disease [2,3,4].
SERCA1 has been largely investigated in structure, domains and functional mechanisms [5,6]. Mutant cDNAs were expressed in cell culture and SERCA1-null mice has been created to investigate the importance of this protein and the implication of the different domains. It has been demonstrated how a single substitution of Arginine 560 (in mouse and rabbit) in domain N leads to a severe alteration of the protein conformation with a substantial reduction of its functionality [7,8].
Our group is working on PMT and interestingly a pathological case, a Dutch cross-breed calf, has shown a natural mutation in ATP2A1 gene leading to a substitution of Arginine 559, corresponding to Arginine 560 in mouse and rabbit. This spontaneous mutation produce a situation similar to the experimental one. Some preliminary histological and biochemical results have been published in a case report [9]. Here we present new histological analysis and early results in the study of the bovine natural mutation in heterologous system
Association of calcineurin with calmodulin, the dihydropyridine receptor and protein kinase A in junctional transverse tubules of rabbit fast-twitch muscle
No full textThe protein to protein interaction of calcineurin with other skeletal muscle proteins was investigate
CaMKII, PP1 and glycogen synthase are present as a functional complex in longitudinal sarcoplasmic reticulum of rabbit fast muscle
No full textJ. MUSCLE RES. CELL MOTIL
The Ca2+-calmodulin dependent protein kinase II system of skeletal muscle sarcoplasmic reticulum
No full textThe association of CaMKII to sarcoplasmic reticulum of skeletal muscle was reviewed. Its role in regulation of glycogen kinase activity was discussed
Colocalization to transverse tubules of dihydropyridine receptor, cAMP-dependent protein kinase and calcineurin and its possible relevance to modulation of excitation-contraction coupling at the triadic junction
No full textIdentification of two ryanodine receptor transcripts in neonatal, slow-, and fast-twitch rabbit skeletal muscles
No full textAnalysis of the primary structure of the rabbit skeletal muscle ryanodine receptor led to the identification of two molecules of 5032 and 5037 residues, respectively. Such a sequence discrepancy is likely to be due to the alternative splicing of a 15 bp exon (1) encoding a 5 amino acid insertion (Ala-Gly-Asp-Ala-Gln) after residue 3479. By using PCR on first strand cDNA, we searched for the 15 base pair insertion in the ryanodine receptor mRNA from adult slow- and fast-twitch skeletal muscle, as well as from fast-muscles, at various stages of post-natal development. All rabbit skeletal muscle mRNAs, regardless of their developmental stage and twitch properties, contain two RYR transcripts, suggesting the coexistence of two RYR isoforms in mammalian skeletal muscl
Clues to calcineurin function in mammalian fast-twitch muscle
No full textIt is believed that brief, high amplitude Ca2+ transients, as found in fast-twitch muscles, are not sufficient to activate the calcineurin (Cn)-dependent signaling pathway involved in regulation of slow myosin and slow sarcoplasmic reticulum Ca2+-ATPase genes (Olson and Williams, Cell 101: 689-692, 2000). The results reported here try to fill the gap between this molecular knowledge, and the still fragmentary pieces of information on a possible different role of calcineurin in the same type of muscles. In the present work calcineurin was determined immunocytochemically by labeling fast- and slow-twitch fibers of representative rabbit muscles with anti-CnB antibodies, and was assessed by western blotting of isolated subcellular fractions. Calcineurin was found to be largely soluble and to be constitutively overexpressed in fast muscle as CnAalpha and CnAbeta isoforms, the latter appearing to be predominant. Particulate calcineurin was not only associated with myofibrils but also with membranes of various origins. Fluorescence microscopy showed that calcineurin was distributed in the same pattern with respect to sarcomeres in both types of fibers, and formed punctate dots spanning the I-Z-I region, rather than being exclusively located at the Z-line, a disposition described for cardiomyocytes (Frey et al., Proc Natl Acad Sci USA 97: 14,632-14,637, 2000). From knowledge that, in mammalian skeletal muscle fibers, junctional triads are located at the A-I band boundary, we explored the distribution of calcineurin between triadic components, after having verified that it was present in very low amounts in dystrophin-enriched sarcolemmal membranes. Our results demonstrate that a small but significant proportion of calcineurin coenriched with transverse tubules (TT), and copurified with the DHPR and with DHPR-associated PKA-AKAP15/18, thus suggesting that it is assembled as a multiprotein complex in the junctional membrane domain of TT. The membrane specificity of this association is further corroborated by the negative evidence for the presence of calcineurin in SR terminal cisternae. Calcineurin was separated from the DHPR and isolated as a AKAP15/18 subcomplex, including beta2 adrenergic receptor, in addition to PKA and calcineurin, following equilibrium centrifugation of detergent extracts on a linear sucrose gradient. We show that the alpha1 subunit skeletal isoform of the DHPR, is a substrate for calcineurin dephosphorylation, after previous phosphorylation by endogenous PKA
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