Mass spectrometric identification of dystrophin, the protein product of the Duchenne muscular dystrophy gene, in distinct muscle surface membranes

Murphy,Sandra; Ohlendieck,Kay

doi:10.3892/ijmm.2017.3082

Mass spectrometric identification of dystrophin, the protein product of the Duchenne muscular dystrophy gene, in distinct muscle surface membranes

Authors:
- Sandra Murphy
- Kay Ohlendieck
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Affiliations: Department of Biology, Maynooth University, National University of Ireland, Maynooth, Co Kildare, Ireland
Published online on: July 27, 2017 https://doi.org/10.3892/ijmm.2017.3082
Pages: 1078-1088
Copyright: © Murphy et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Supramolecular membrane complexes of low abundance are difficult to study by routine bioanalytical techniques. The plasmalemmal complex consisting of sarcoglycans, dystroglycans, dystrobrevins and syntrophins, which is closely associated with the membrane cytoskeletal protein dystrophin, represents such a high‑molecular‑mass protein assembly in skeletal muscles. The almost complete loss of the dystrophin isoform Dp427‑M and concomitant reduction in the dystrophin‑associated glycoprotein complex is the underlying cause of the highly progressive neuromuscular disorder named Duchenne muscular dystrophy. This gives the detailed characterization of the dystrophin complex considerable pathophysiological importance. In order to carry out a comprehensive mass spectrometric identification of the dystrophin‑glycoprotein complex, in this study, we used extensive subcellular fractionation and enrichment procedures prior to subproteomic analysis. Mass spectrometry identified high levels of full‑length dystrophin isoform Dp427‑M, α/β‑dystroglycans, α/β/γ/δ‑sarcoglycans, α1/β1/β2‑syntrophins and α/β‑dystrobrevins in highly purified sarcolemma vesicles. By contrast, lower levels were detected in transverse tubules and no components of the dystrophin complex were identified in triads. For comparative purposes, the presence of organellar marker proteins was studied in crude surface membrane preparations vs. enriched fractions from the sarcolemma, transverse tubules and triad junctions using gradient gel electrophoresis and on‑membrane digestion. This involved the subproteomic assessment of various ion‑regulatory proteins and excitation‑contraction coupling components. The comparative profiling of skeletal muscle fractions established a relatively restricted subcellular localization of the dystrophin‑glycoprotein complex in the muscle fibre periphery by proteomic means and clearly demonstrated the absence of dystrophin from triad junctions by sensitive mass spectrometric analysis.

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1	Cifani P and Kentsis A: Towards comprehensive and quantitative proteomics for diagnosis and therapy of human disease. Proteomics. 17:Jan;2017.Epub ahead of print. View Article : Google Scholar
2	Angel TE, Aryal UK, Hengel SM, Baker ES, Kelly RT, Robinson EW and Smith RD: Mass spectrometry-based proteomics: Existing capabilities and future directions. Chem Soc Rev. 41:3912–3928. 2012. View Article : Google Scholar : PubMed/NCBI
3	Van Riper SK, de Jong EP, Carlis JV and Griffin TJ: Mass spectrometry-based proteomics: Basic principles and emerging technologies and directions. Adv Exp Med Biol. 990:1–35. 2013. View Article : Google Scholar : PubMed/NCBI
4	Zhang Z, Wu S, Stenoien DL and Paša-Tolić L: High-throughput proteomics. Annu Rev Anal Chem (Palo Alto, Calif). 7:427–454. 2014. View Article : Google Scholar
5	Allen DG, Whitehead NP and Froehner SC: Absence of dystrophin disrupts skeletal muscle signaling: Roles of Ca²⁺, reactive oxygen species, and nitric oxide in the development of muscular dystrophy. Physiol Rev. 96:253–305. 2016. View Article : Google Scholar
6	Holland A, Murphy S, Dowling P and Ohlendieck K: Pathoproteomic profiling of the skeletal muscle matrisome in dystrophinopathy associated myofibrosis. Proteomics. 16:345–366. 2016. View Article : Google Scholar
7	Ohlendieck K and Swandulla D: Molecular pathogenesis of Duchenne muscular dystrophy-related fibrosis. Pathologe. 38:21–29. 2017.In German. View Article : Google Scholar : PubMed/NCBI
8	Murphy S and Ohlendieck K: The biochemical and mass spectrometric profiling of the dystrophin complexome from skeletal muscle. Comput Struct Biotechnol J. 14:20–27. 2015. View Article : Google Scholar
9	Fuller HR, Graham LC, Llavero Hurtado M and Wishart TM: Understanding the molecular consequences of inherited muscular dystrophies: Advancements through proteomic experimentation. Expert Rev Proteomics. 13:659–671. 2016. View Article : Google Scholar : PubMed/NCBI
10	Guiraud S, Aartsma-Rus A, Vieira NM, Davies KE, van Ommen GJ and Kunkel LM: The pathogenesis and therapy of muscular dystrophies. Annu Rev Genomics Hum Genet. 16:281–308. 2015. View Article : Google Scholar : PubMed/NCBI
11	Doran P, Martin G, Dowling P, Jockusch H and Ohlendieck K: Proteome analysis of the dystrophin-deficient MDX diaphragm reveals a drastic increase in the heat shock protein cvHSP. Proteomics. 6:4610–4621. 2006. View Article : Google Scholar : PubMed/NCBI
12	Rayavarapu S, Coley W, Cakir E, Jahnke V, Takeda S, Aoki Y, Grodish-Dressman H, Jaiswal JK, Hoffman EP, Brown KJ, et al: Identification of disease specific pathways using in vivo SILAC proteomics in dystrophin deficient mdx mouse. Mol Cell Proteomics. 12:1061–1073. 2013. View Article : Google Scholar : PubMed/NCBI
13	Holland A, Henry M, Meleady P, Winkler CK, Krautwald M, Brinkmeier H and Ohlendieck K: Comparative label-free mass spectrometric analysis of mildly versus severely affected mdx mouse skeletal muscles identifies Annexin, lamin, and Vimentin as universal dystrophic markers. Molecules. 20:11317–11344. 2015. View Article : Google Scholar : PubMed/NCBI
14	Holland A, Dowling P, Meleady P, Henry M, Zweyer M, Mundegar RR, Swandulla D and Ohlendieck K: Label-free mass spectrometric analysis of the mdx-4cv diaphragm identifies the matricellular protein periostin as a potential factor involved in dystrophinopathy-related fibrosis. Proteomics. 15:2318–2331. 2015. View Article : Google Scholar : PubMed/NCBI
15	Holland A, Carberry S and Ohlendieck K: Proteomics of the dystrophin-glycoprotein complex and dystrophinopathy. Curr Protein Pept Sci. 14:680–697. 2013. View Article : Google Scholar : PubMed/NCBI
16	Dowling P, Holland A and Ohlendieck K: Mass spectrometry-based identification of muscle-associated and muscle-derived proteomic biomarkers of dystrophinopathies. J Neuromuscul Dis. 1:15–40. 2014.PubMed/NCBI
17	Murphy S, Dowling P and Ohlendieck K: Comparative skeletal muscle proteomics using two-dimensional gel electrophoresis. Proteomes. 4:272016. View Article : Google Scholar
18	Lewis C and Ohlendieck K: Mass spectrometric identification of dystrophin isoform Dp427 by on-membrane digestion of sarcolemma from skeletal muscle. Anal Biochem. 404:197–203. 2010. View Article : Google Scholar : PubMed/NCBI
19	Yoon JH, Johnson E, Xu R, Martin LT, Martin PT and Montanaro F: Comparative proteomic profiling of dystroglycan-associated proteins in wild type, mdx, and Galgt2 transgenic mouse skeletal muscle. J Proteome Res. 11:4413–4424. 2012. View Article : Google Scholar : PubMed/NCBI
20	Murphy S, Henry M, Meleady P, Zweyer M, Mundegar RR, Swandulla D and Ohlendieck K: Simultaneous pathoproteomic evaluation of the dystrophin-glycoprotein complex and secondary changes in the mdx-4cv mouse model of Duchenne muscular dystrophy. Biology (Basel). 4:397–423. 2015.
21	Murphy S, Zweyer M, Mundegar RR, Henry M, Meleady P, Swandulla D and Ohlendieck K: Concurrent label-free mass spectrometric analysis of dystrophin isoform Dp427 and the myofibrosis marker collagen in crude extracts from mdx-4cv skeletal muscles. Proteomes. 3:298–327. 2015. View Article : Google Scholar : PubMed/NCBI
22	Turk R, Hsiao JJ, Smits MM, Ng BH, Pospisil TC, Jones KS, Campbell KP and Wright ME: Molecular signatures of membrane protein complexes underlying muscular dystrophy. Mol Cell Proteomics. 15:2169–2185. 2016. View Article : Google Scholar : PubMed/NCBI
23	Bonilla E, Samitt CE, Miranda AF, Hays AP, Salviati G, DiMauro S, Kunkel LM, Hoffman EP and Rowland LP: Duchenne muscular dystrophy: Deficiency of dystrophin at the muscle cell surface. Cell. 54:447–452. 1988. View Article : Google Scholar : PubMed/NCBI
24	Hoffman EP, Knudson CM, Campbell KP and Kunkel LM: Subcellular fractionation of dystrophin to the triads of skeletal muscle. Nature. 330:754–758. 1987. View Article : Google Scholar : PubMed/NCBI
25	Knudson CM, Hoffman EP, Kahl SD, Kunkel LM and Campbell KP: Evidence for the association of dystrophin with the transverse tubular system in skeletal muscle. J Biol Chem. 263:8480–8484. 1988.PubMed/NCBI
26	Salviati G, Betto R, Ceoldo S, Biasia E, Bonilla E, Miranda AF and Dimauro S: Cell fractionation studies indicate that dystrophin is a protein of surface membranes of skeletal muscle. Biochem J. 258:837–841. 1989. View Article : Google Scholar : PubMed/NCBI
27	Ohlendieck K, Ervasti JM, Snook JB and Campbell KP: Dystrophin-glycoprotein complex is highly enriched in isolated skeletal muscle sarcolemma. J Cell Biol. 112:135–148. 1991. View Article : Google Scholar : PubMed/NCBI
28	Ohlendieck K and Campbell KP: Dystrophin constitutes 5% of membrane cytoskeleton in skeletal muscle. FEBS Lett. 283:230–234. 1991. View Article : Google Scholar : PubMed/NCBI
29	Ohlendieck K and Campbell KP: Dystrophin-associated proteins are greatly reduced in skeletal muscle from mdx mice. J Cell Biol. 115:1685–1694. 1991. View Article : Google Scholar : PubMed/NCBI
30	Ohlendieck K, Matsumura K, Ionasescu VV, Towbin JA, Bosch EP, Weinstein SL, Sernett SW and Campbell KP: Duchenne muscular dystrophy: Deficiency of dystrophin-associated proteins in the sarcolemma. Neurology. 43:795–800. 1993. View Article : Google Scholar : PubMed/NCBI
31	Dowling P, Lohan J and Ohlendieck K: Comparative analysis of Dp427-deficient mdx tissues shows that the milder dystrophic phenotype of extraocular and toe muscle fibres is associated with a persistent expression of beta-dystroglycan. Eur J Cell Biol. 82:222–230. 2003. View Article : Google Scholar : PubMed/NCBI
32	Cluchague N, Moreau C, Rocher C, Pottier S, Leray G, Cherel Y and Le Rumeur E: beta-Dystroglycan can be revealed in microsomes from mdx mouse muscle by detergent treatment. FEBS Lett. 572:216–220. 2004. View Article : Google Scholar : PubMed/NCBI
33	Daval S, Rocher C, Cherel Y and Le Rumeur E: Several dystrophin-glycoprotein complex members are present in crude surface membranes but they are sodium dodecyl sulphate invisible in KCl-washed microsomes from mdx mouse muscle. Cell Mol Biol Lett. 15:134–152. 2010. View Article : Google Scholar
34	Ohlendieck K: On-membrane digestion technology for muscle proteomics. J Membr Sep Technol. 2:1–12. 2013.
35	Ohlendieck K: Organelle proteomics in skeletal muscle biology. J Integr OMICS. 2:27–38. 2012. View Article : Google Scholar
36	Zubrzycka-Gaarn EE, Bulman DE, Karpati G, Burghes AH, Belfall B, Klamut HJ, Talbot J, Hodges RS, Ray PN and Worton RG: The Duchenne muscular dystrophy gene product is localized in sarcolemma of human skeletal muscle. Nature. 333:466–469. 1988. View Article : Google Scholar : PubMed/NCBI
37	Watkins SC, Hoffman EP, Slayter HS and Kunkel LM: Immunoelectron microscopic localization of dystrophin in myofibres. Nature. 333:863–866. 1988. View Article : Google Scholar : PubMed/NCBI
38	Cullen MJ, Walsh J, Nicholson LV and Harris JB: Ultrastructural localization of dystrophin in human muscle by using gold immunolabelling. Proc R Soc Lond B Biol Sci. 240:197–210. 1990. View Article : Google Scholar : PubMed/NCBI
39	Carpenter S, Karpati G, Zubrzycka-Gaarn E, Bulman DE, Ray PN and Worton RG: Dystrophin is localized to the plasma membrane of human skeletal muscle fibers by electron-microscopic cytochemical study. Muscle Nerve. 13:376–380. 1990. View Article : Google Scholar : PubMed/NCBI
40	Staunton L, Jockusch H, Wiegand C, Albrecht T and Ohlendieck K: Identification of secondary effects of hyperexcit-ability by proteomic profiling of myotonic mouse muscle. Mol Biosyst. 7:2480–2489. 2011. View Article : Google Scholar : PubMed/NCBI
41	Murray BE and Ohlendieck K: Cross-linking analysis of the ryanodine receptor and alpha1-dihydropyridine receptor in rabbit skeletal muscle triads. Biochem J. 324:689–696. 1997. View Article : Google Scholar : PubMed/NCBI
42	Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 72:248–254. 1976. View Article : Google Scholar : PubMed/NCBI
43	Rosemblatt M, Hidalgo C, Vergara C and Ikemoto N: Immunological and biochemical properties of transverse tubule membranes isolated from rabbit skeletal muscle. J Biol Chem. 56:8140–8148. 1981.
44	Sharp AH, Imagawa T, Leung AT and Campbell KP: Identification and characterization of the dihydropyridine-binding subunit of the skeletal muscle dihydropyridine receptor. J Biol Chem. 262:12309–12315. 1987.PubMed/NCBI
45	Muñoz P, Rosemblatt M, Testar X, Palacín M and Zorzano A: Isola tion and characterization of distinct domains of sarcolemma and T-tubules from rat skeletal muscle. Biochem J. 307:273–280. 1995. View Article : Google Scholar
46	Staunton L and Ohlendieck K: Mass spectrometric characterization of the sarcoplasmic reticulum from rabbit skeletal muscle by on-membrane digestion. Protein Pept Lett. 19:252–263. 2012. View Article : Google Scholar
47	Ohlendieck K: Characterisation of the dystrophin-related protein utrophin in highly purified skeletal muscle sarcolemma vesicles. Biochim Biophys Acta. 1283:215–222. 1996. View Article : Google Scholar : PubMed/NCBI
48	Vretblad P: Purification of lectins by biospecific affinity chromatography. Biochim Biophys Acta. 434:169–176. 1976. View Article : Google Scholar : PubMed/NCBI
49	Luque-Garcia JL, Zhou G, Sun TT and Neubert TA: Use of nitrocellulose membranes for protein characterization by matrix-assisted laser desorption/ionization mass spectrometry. Anal Chem. 78:5102–5108. 2006. View Article : Google Scholar : PubMed/NCBI
50	Luque-Garcia JL, Zhou G, Spellman DS, Sun TT and Neubert TA: Analysis of electroblotted proteins by mass spectrometry: Protein identification after western blotting. Mol Cell Proteomics. 7:308–314. 2008. View Article : Google Scholar
51	Luque-Garcia JL and Neubert TA: On-membrane tryptic digestion of proteins for mass spectrometry analysis. Methods Mol Biol. 536:331–341. 2009. View Article : Google Scholar : PubMed/NCBI
52	Shevchenko A, Tomas H, Havlis J, Olsen JV and Mann M: In-gel digestion for mass spectrometric characterization of proteins and proteomes. Nat Protoc. 1:2856–2860. 2006. View Article : Google Scholar
53	Murphy S, Dowling P, Zweyer M, Mundegar RR, Henry M, Meleady P, Swandulla D and Ohlendieck K: Proteomic analysis of dystrophin deficiency and associated changes in the aged mdx-4cv heart model of dystrophinopathy-related cardiomyopathy. J Proteomics. 145:24–36. 2016. View Article : Google Scholar : PubMed/NCBI
54	Liu Y, Bouhenni RA, Dufresne CP, Semba RD and Edward DP: Differential expression of vitreous proteins in young and mature New Zealand white rabbits. PLoS One. 11:e01535602016. View Article : Google Scholar : PubMed/NCBI
55	Ryan M, Butler-Browne G, Erzen I, Mouly V, Thornell LE, Wernig A and Ohlendieck K: Persistent expression of the alpha1S-dihydropyridine receptor in aged human skeletal muscle: Implications for the excitation-contraction uncoupling hypothesis of sarcopenia. Int J Mol Med. 11:425–434. 2003.PubMed/NCBI
56	Glover L, Heffron JJ and Ohlendieck K: Increased sensitivity of the ryanodine receptor to halothane-induced oligomerization in malignant hyperthermia-susceptible human skeletal muscle. J Appl Physiol 1985. 96:11–18. 2004. View Article : Google Scholar
57	Staunton L, Zweyer M, Swandulla D and Ohlendieck K: Mass spectrometry-based proteomic analysis of middle-aged vs. aged vastus lateralis reveals increased levels of carbonic anhydrase isoform 3 in senescent human skeletal muscle. Int J Mol Med. 30:723–733. 2012. View Article : Google Scholar : PubMed/NCBI
58	Chartier A, Klein P, Pierson S, Barbezier N, Gidaro T, Casas F, Carberry S, Dowling P, Maynadier L, Bellec M, et al: Mitochondrial dysfunction reveals the role of mRNA poly(A) tail regulation in oculopharyngeal muscular dystrophy pathogenesis. PLoS Genet. 11:e10050922015. View Article : Google Scholar : PubMed/NCBI
59	Drissi R, Dubois ML and Boisvert FM: Proteomics methods for subcellular proteome analysis. FEBS J. 280:5626–5634. 2013. View Article : Google Scholar : PubMed/NCBI
60	Breckels LM, Gatto L, Christoforou A, Groen AJ, Lilley KS and Trotter MW: The effect of organelle discovery upon sub-cellular protein localisation. J Proteomics. 88:129–140. 2013. View Article : Google Scholar : PubMed/NCBI
61	Mueller SJ, Hoernstein SN and Reski R: Approaches to characterize organelle, compartment, or structure purity. Methods Mol Biol. 1511:13–28. 2017. View Article : Google Scholar
62	Larance M and Lamond AI: Multidimensional proteomics for cell biology. Nat Rev Mol Cell Biol. 16:269–280. 2015. View Article : Google Scholar : PubMed/NCBI
63	Lefort N, Yi Z, Bowen B, Glancy B, De Filippis EA, Mapes R, Hwang H, Flynn CR, Willis WT, Civitarese A, et al: Proteome profile of functional mitochondria from human skeletal muscle using one-dimensional gel electrophoresis and HPLC-ESI-MS/MS. J Proteomics. 72:1046–1060. 2009. View Article : Google Scholar : PubMed/NCBI
64	Lombardi A, Silvestri E, Cioffi F, Senese R, Lanni A, Goglia F, de Lange P and Moreno M: Defining the transcriptomic and proteomic profiles of rat ageing skeletal muscle by the use of a cDNA array, 2D- and Blue native-PAGE approach. J Proteomics. 72:708–721. 2009. View Article : Google Scholar : PubMed/NCBI
65	Ferreira R, Vitorino R, Alves RM, Appell HJ, Powers SK, Duarte JA and Amado F: Subsarcolemmal and intermyofibrillar mitochondria proteome differences disclose functional specializations in skeletal muscle. Proteomics. 10:3142–3154. 2010. View Article : Google Scholar : PubMed/NCBI
66	Liu Z, Du X, Deng J, Gu M, Hu H, Gui M, Yin CC and Chang Z: The interactions between mitochondria and sarcoplasmic reticulum and the proteome characterization of mitochondrion-associated membrane from rabbit skeletal muscle. Proteomics. 15:2701–2704. 2015. View Article : Google Scholar : PubMed/NCBI
67	Liu Z, Du X, Yin C and Chang Z: Shotgun proteomic analysis of sarcoplasmic reticulum preparations from rabbit skeletal muscle. Proteomics. 13:2335–2338. 2013. View Article : Google Scholar : PubMed/NCBI
68	Vitorino R, Ferreira R, Neuparth M, Guedes S, Williams J, Tomer KB, Domingues PM, Appell HJ, Duarte JA and Amado M: Subcellular proteomics of mice gastrocnemius and soleus muscles. Anal Biochem. 366:156–169. 2007. View Article : Google Scholar : PubMed/NCBI
69	Toigo M, Donohoe S, Sperrazzo G, Jarrold B, Wang F, Hinkle R, Dolan E, Isfort RJ and Aebersold R: ICAT-MS-MS time course analysis of atrophying mouse skeletal muscle cytosolic subproteome. Mol Biosyst. 1:229–241. 2005. View Article : Google Scholar
70	Maughan DW, Henkin JA and Vigoreaux JO: Concentrations of glycolytic enzymes and other cytosolic proteins in the diffusible fraction of a vertebrate muscle proteome. Mol Cell Proteomics. 4:1541–1549. 2005. View Article : Google Scholar : PubMed/NCBI
71	Ohlendieck K: Proteomics of skeletal muscle glycolysis. Biochim Biophys Acta. 1804:2089–2101. 2010. View Article : Google Scholar : PubMed/NCBI
72	Gannon J, Doran P, Kirwan A and Ohlendieck K: Drastic increase of myosin light chain MLC-2 in senescent skeletal muscle indicates fast-to-slow fibre transition in sarcopenia of old age. Eur J Cell Biol. 88:685–700. 2009. View Article : Google Scholar : PubMed/NCBI
73	Holland A and Ohlendieck K: Proteomic profiling of the contractile apparatus from skeletal muscle. Expert Rev Proteomics. 10:239–257. 2013. View Article : Google Scholar : PubMed/NCBI
74	Ohlendieck K: Towards an understanding of the dystrophin-glycoprotein complex: Linkage between the extracellular matrix and the membrane cytoskeleton in muscle fibers. Eur J Cell Biol. 69:1–10. 1996.PubMed/NCBI
75	Murray BE, Froemming GR, Maguire PB and Ohlendieck K: Excitation-contraction-relaxation cycle: Role of Ca²⁺-regulatory membrane proteins in normal, stimulated and pathological skeletal muscle (Review). Int J Mol Med. 1:677–687. 1998.PubMed/NCBI