Name: quantitative analysis of chromatin proteomes in disease
Uploaded: 2012-12-28T12:00:00
Description: Watch this Scientific Journal Video about Quantitative Analysis of Chromatin Proteomes in Disease at JoVE.com

De Vondriska lab wordt ondersteund door subsidies van de National Heart, Lung and Blood Institute van de NIH en de Laubisch Endowment aan de UCLA. EM is ontvanger van de Jennifer S. Buchwald Graduate Fellowship voor de Fysiologie aan de UCLA, HC is de ontvanger van een American Heart Association Pre-doctorale beurs, MP is de ontvanger van een NIH Ruth Kirschstein post-doctorale beurs, en SF is de ontvanger van een NIH K99 Award.

De experimentele workflow bevat zeven belangrijke stappen (Figuur 1). Voor alle werkzaamheden waarbij monsters die zullen worden uitgevoerd op de massaspectrometer, moet er worden dragen een laboratoriumjas, handschoenen en haarnetje en zorg om besmetting tegen stof en persoonlijke vergieten van keratine te voorkomen. 1. Hart Homogenisatie en nucleaire Isolatie Muis harten gehomogeniseerd en een intacte kernen pellet wordt geïsoleerd (Figu...

<ol>
	<li>Li, F., Wang, X., Capasso, J. M., Gerdes, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+transition+of+cardiac+myocytes+from+hyperplasia+to+hypertrophy+during+postnatal+development.">Rapid transition of cardiac myocytes from hyperplasia to hypertrophy during postnatal development.</a> J Mol Cell Cardiol. 28, 1737-1746 (1996).</li><li>Rumyantsev, P. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interrelations+of+the+proliferation+and+differentiation+processes+during+cardiact+myogenesis+and+regeneration.">Interrelations of the proliferation and differentiation processes during cardiact myogenesis and regeneration.</a> Int Rev. Cytol. 51, 186-273 (1977).</li><li>Olson, E. N., Schneider, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sizing+up+the+heart:+development+redux+in+disease.">Sizing up the heart: development redux in disease.</a> Genes Dev. 17, 1937-1956 (2003).</li><li>Molkentin, J. D., Dorn, G. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cytoplasmic+signaling+pathways+that+regulate+cardiac+hypertrophy.">Cytoplasmic signaling pathways that regulate cardiac hypertrophy.</a> Annu Rev. Physiol. 63, 391-426 (2001).</li><li>Fishman, M. C., Chien, K. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fashioning+the+vertebrate+heart:+earliest+embryonic+decisions.">Fashioning the vertebrate heart: earliest embryonic decisions.</a> Development. 124, 2099-2117 (1997).</li><li>Rajabi, M., Kassiotis, C., Razeghi, P., Taegtmeyer, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Return+to+the+fetal+gene+program+protects+the+stressed+heart:+a+strong+hypothesis.">Return to the fetal gene program protects the stressed heart: a strong hypothesis.</a> Heart Fail Rev. 12, 331-343 (2007).</li><li>Roger, V. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heart+disease+and+stroke+statistics--2011+update:+a+report+from+the+American+Heart+Association.">Heart disease and stroke statistics--2011 update: a report from the American Heart Association.</a> Circulation. 123, e18-e209 (2011).</li><li>Schirmer, E. C., Florens, L., Guan, T., Yates, J. R., Gerace, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nuclear+membrane+proteins+with+potential+disease+links+found+by+subtractive+proteomics.">Nuclear membrane proteins with potential disease links found by subtractive proteomics.</a> Science. 301, 1380-1382 (2003).</li><li>Lambert, J. P., Mitchell, L., Rudner, A., Baetz, K., Figeys, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+proteomics+approach+for+the+discovery+of+chromatin-associated+protein+networks.">A novel proteomics approach for the discovery of chromatin-associated protein networks.</a> Mol. Cell Proteomics. 8, 870-882 (2009).</li><li>Malik, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell-specific+and+lamin-dependent+targeting+of+novel+transmembrane+proteins+in+the+nuclear+envelope.">Cell-specific and lamin-dependent targeting of novel transmembrane proteins in the nuclear envelope.</a> Cell Mol Life Sci. 67, 1353-1369 (2010).</li><li>Tweedie-Cullen, R. Y., Reck, J. M., Mansuy, I. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comprehensive+mapping+of+post-translational+modifications+on+synaptic,+nuclear,+and+histone+proteins+in+the+adult+mouse+brain.">Comprehensive mapping of post-translational modifications on synaptic, nuclear, and histone proteins in the adult mouse brain.</a> J. Proteome Res. 8, 4966-4982 (2009).</li><li>Chu, D. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sperm+chromatin+proteomics+identifies+evolutionarily+conserved+fertility+factors.">Sperm chromatin proteomics identifies evolutionarily conserved fertility factors.</a> Nature. 443, 101-105 (2006).</li><li>Uchiyama, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proteome+analysis+of+human+metaphase+chromosomes.">Proteome analysis of human metaphase chromosomes.</a> J. Biol. Chem. 280, 16994-17004 (2005).</li><li>Franklin, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Specialized+compartments+of+cardiac+nuclei+exhibit+distinct+proteomic+anatomy.">Specialized compartments of cardiac nuclei exhibit distinct proteomic anatomy.</a> Mol. Cell Proteomics. 10, (2011).</li><li>Kislinger, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+survey+of+organ+and+organelle+protein+expression+in+mouse:+combined+proteomic+and+transcriptomic+profiling.">Global survey of organ and organelle protein expression in mouse: combined proteomic and transcriptomic profiling.</a> Cell. 125, 173-186 (2006).</li><li>Moore, D. H., Ruska, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electron+microscope+study+of+mammalian+cardiac+muscle+cells.">Electron microscope study of mammalian cardiac muscle cells.</a> J. Biophys Biochem. Cytol. 3, 261-268 (1957).</li><li>Anversa, P., Vitali-Mazza, L., Visioli, O., Marchetti, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+cardiac+hypertrophy:+a+quantitative+ultrastructural+study+in+the+compensatory+stage.">Experimental cardiac hypertrophy: a quantitative ultrastructural study in the compensatory stage.</a> J. Mol. Cell Cardiol. 3, 213-227 (1971).</li><li>Franklin, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Specialized+compartments+of+cardiac+nuclei+exhibit+distinct+proteomic+anatomy.">Specialized compartments of cardiac nuclei exhibit distinct proteomic anatomy.</a> Mol Cell. Proteomics. 10, (2011).</li><li>Paulsson, A. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Post-translational+regulation+of+calsarcin-1+during+pressure+overload-induced+cardiac+hypertrophy.">Post-translational regulation of calsarcin-1 during pressure overload-induced cardiac hypertrophy.</a> Journal of molecular and cellular cardiology. 48, 1206-1214 (2010).</li><li>Franklin, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+analysis+of+the+chromatin+proteome+in+disease+reveals+remodeling+principles+and+identifies+high+mobility+group+protein+b2+as+a+regulator+of+hypertrophic+growth.">Quantitative analysis of the chromatin proteome in disease reveals remodeling principles and identifies high mobility group protein b2 as a regulator of hypertrophic growth.</a> Mol Cell Proteomics. 11, (2012).</li><li>Gramolini, A. O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+proteomics+profiling+of+a+phospholamban+mutant+mouse+model+of+dilated+cardiomyopathy+reveals+progressive+intracellular+stress+responses.">Comparative proteomics profiling of a phospholamban mutant mouse model of dilated cardiomyopathy reveals progressive intracellular stress responses.</a> Mol Cell Proteomics. 7, 519-533 (2008).</li><li>Singh, H., Warburton, S., Vondriska, T. M., Khakh, B. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proteomics+to+Identify+Proteins+Interacting+with+P2X2+Ligand-Gated+Cation+Channels.">Proteomics to Identify Proteins Interacting with P2X2 Ligand-Gated Cation Channels.</a> J. Vis. Exp. (27), e1178 (2009).</li><li>Park, S. K., Venable, J. D., Xu, T., Yates, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+quantitative+analysis+software+tool+for+mass+spectrometry-based+proteomics.">A quantitative analysis software tool for mass spectrometry-based proteomics.</a> Nat. Methods. 5, 319-322 (2008).</li><li>Lomenick, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Target+identification+using+drug+affinity+responsive+target+stability+(DARTS).">Target identification using drug affinity responsive target stability (DARTS).</a> Proc. Natl. Acad. Sci. U. S. A. 106, 21984-21989 (2009).</li><li>Kamleh, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Metabolomic+profiling+using+Orbitrap+Fourier+transform+mass+spectrometry+with+hydrophilic+interaction+chromatography:+a+method+with+wide+applicability+to+analysis+of+biomolecules.">Metabolomic profiling using Orbitrap Fourier transform mass spectrometry with hydrophilic interaction chromatography: a method with wide applicability to analysis of biomolecules.</a> Rapid Commun. Mass Spectrom. 22, 1912-1918 (2008).</li><li>Searle, B. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scaffold:+a+bioinformatic+tool+for+validating+MS/MS-based+proteomic+studies.">Scaffold: a bioinformatic tool for validating MS/MS-based proteomic studies.</a> Proteomics. 10, 1265-1269 (2010).</li><li>Murray, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Basic+Proteins+of+Cell+Nuclei.">The Basic Proteins of Cell Nuclei.</a> Annu Rev. Biochem. 34, 209-246 (1965).</li><li>Johns, E. W., Phillips, D. M., Simson, P., Butler, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improved+fractionations+of+arginine-rich+histones+from+calf+thymus.">Improved fractionations of arginine-rich histones from calf thymus.</a> Biochem. J. 77, 631-636 (1960).</li><li>Rodriguez-Collazo, P., Leuba, S. H., Zlatanova, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Robust+methods+for+purification+of+histones+from+cultured+mammalian+cells+with+the+preservation+of+their+native+modifications.">Robust methods for purification of histones from cultured mammalian cells with the preservation of their native modifications.</a> Nucleic Acids Res. 37, e81 (2009).</li><li>Kuehl, L., Salmond, B., Tran, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Concentrations+of+high-mobility-group+proteins+in+the+nucleus+and+cytoplasm+of+several+rat+tissues.">Concentrations of high-mobility-group proteins in the nucleus and cytoplasm of several rat tissues.</a> J. Cell Biol. 99, 648-654 (1984).</li><li>Gronow, M., Griffiths, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+isolation+and+separation+of+the+non-histone+proteins+of+rat+liver+nuclei.">Rapid isolation and separation of the non-histone proteins of rat liver nuclei.</a> FEBS Lett. 15, 340-344 (1971).</li><li>Mischke, B. G., Ward, O. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+tissue+specificity+of+chromatin-associated+proteins+in+Vicia+faba.">Isolation and tissue specificity of chromatin-associated proteins in Vicia faba.</a> Can J. Biochem. 53, 91-95 (1975).</li><li>Andersen, J. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Directed+proteomic+analysis+of+the+human+nucleolus.">Directed proteomic analysis of the human nucleolus.</a> Curr. Biol. 12, 1-11 (2002).</li><li>Zhao, Y., Jensen, O. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modification-specific+proteomics:+strategies+for+characterization+of+post-translational+modifications+using+enrichment+techniques.">Modification-specific proteomics: strategies for characterization of post-translational modifications using enrichment techniques.</a> Proteomics. 9, 4632-4641 (2009).</li><li>Ahrne, E., Muller, M., Lisacek, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Unrestricted+identification+of+modified+proteins+using+MS/MS.">Unrestricted identification of modified proteins using MS/MS.</a> Proteomics. 10, 671-686 (2010).</li><li>Young, N. L., Plazas-Mayorca, M. D., Garcia, B. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Systems-wide+proteomic+characterization+of+combinatorial+post-translational+modification+patterns.">Systems-wide proteomic characterization of combinatorial post-translational modification patterns.</a> Expert Rev. Proteomics. 7, 79-92 (2010).</li><li>Guthals, A., Bandeira, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Peptide+identification+by+tandem+mass+spectrometry+with+alternate+fragmentation+modes.">Peptide identification by tandem mass spectrometry with alternate fragmentation modes.</a> Mol. Cell Proteomics. , (2012).</li><li>Wu, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+protease+for+'middle-down'+proteomics.">A protease for 'middle-down' proteomics.</a> Nat. Methods. , (2012).</li><li>Tran, J. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mapping+intact+protein+isoforms+in+discovery+mode+using+top-down+proteomics.">Mapping intact protein isoforms in discovery mode using top-down proteomics.</a> Nature. 480, 254-258 (2011).</li></ol>

Twee methoden voor nucleaire isolatie eerder zijn beoordeeld: 27 men de techniek van Behrens homogeniseren gelyofiliseerd weefsel in een niet-waterig oplosmiddel en de tweede een wijziging waarvan we hier homogeniseren van weefsel in een waterige sucrose / zoutoplossing gevolgd door differentiële of dichtheid-gradiënt centrifugatie. Subfractionation van de kernen door zure extractie op weefselmonsters is een belangrijk instrument voor het bestuderen van chromatine die is sinds 1...

<table border="1"><tbody><tr><td> Naam van het reagens </td><td> Vennootschap </td><td> Catalogusnummer </td></tr><tr><td> Dulbeco Modified Eagle Medium </td><td> Invitrogen </td><td> 11965 </td></tr><tr><td> Protease pellet </td><td> Roche </td><td> 04 693 159 001 </td></tr><tr><td> 100 um zeef </td><td> BD Falcon </td><td> 352360 </td></tr><tr><td> Ultracut ultramicrotoom </td><td> Reichert </td><td></td></tr><tr><td> 100CX Transmissie Elektronen Microscoop </td><td> JEOL USA, Inc </td><td></td></tr><tr><td> Wielewaal </td><td> BioRad </td><td> 161-0496 </td></tr><tr><td> Histon H2A antilichaam </td><td> Santa Cruz </td><td> sc-8648 </td></tr><tr><td> Nucleoporin p62 antilichaam </td><td> BD Biosciences </td><td> 610498 </td></tr><tr><td> Adenine nucleotij transporter antilichaam </td><td> Santa Cruz </td><td> sc-9299 </td></tr><tr><td> BiP antilichaam </td><td> Santa Cruz </td><td> sc-1050 </td></tr><tr><td> Tubuline antilichaam </td><td> Sigma </td><td> T1568 </td></tr><tr><td> Histon H3 antilichaam </td><td> Abcam </td><td> ab1791 </td></tr><tr><td> Fibrillarin antilichaam </td><td> Cell Signaling </td><td> C12C3 </td></tr><tr><td> SNRP70 antilichaam </td><td> Abcam </td><td> ab51266 </td></tr><tr><td> E2F-1 antilichaam </td><td> Thermo Fisher </td><td> MS-879 </td></tr><tr><td> Retinoblastoom antilichaam </td><td> BD Biosciences </td><td> 554136 </td></tr><tr><td> Hypoxie induceerbare factor-1 antilichaam </td><td> Novus Biologicals </td><td> NB100-469 </td></tr><tr><td> BCA eiwitbepaling </td><td> Thermo Scientific </td><td> 23227 </td></tr><tr><td> Omgekeerde fasekolom </td><td> Nieuwe Objectief </td><td> PFC7515-B14-10 </td></tr><tr><td> BioWorks Browser </td><td> Thermo Scientific </td><td></td></tr></tbody></table>

quantitative analysis of chromatin proteomes in disease

In de kern bevinden de proteomen waarvan de functies zijn het meest nauw verbonden met de genregulatie. Volwassen zoogdier cardiomyocyte kernen zijn uniek vanwege het hoge percentage binucleaire cellen, de overwegend een heterochromatine toestand van het DNA en de niet-delende aard van de cardiomyocyt waarin volwassen kernen maakt in een permanente staat van interfase. Twee Transcriptionele regulatie tijdens de ontwikkeling en ziekte zijn goed bestudeerd in dit orgaan, 3-5 maar wat blijft relatief onontdekt is de rol van de nucleaire eiwitten die verantwoordelijk zijn voor DNA verpakking en expressie, en hoe deze eiwitten veranderingen in transcriptie&#39;s die tijdens ziekte te bestrijden. 6 In de ontwikkelde wereld, hart-en vaatziekten is de nummer een doodsoorzaak voor zowel mannen als vrouwen. 7 Inzicht in hoe nucleaire eiwitten samen te werken om de progressie van deze ziekte te reguleren is van cruciaal belang voor het bevorderen van de huidige behandeling oOPTIES. Massaspectrometrie is de ideale tool voor het aanpakken van deze vragen omdat het zorgt voor een onpartijdige annotatie van de nucleaire proteoom en de relatieve kwantificatie voor de manier waarop de overvloed van deze eiwitten verandert met de ziekte. Hoewel er verschillende proteomics studies voor zoogdieren nucleaire eiwit-complexen, 8-13 tot voor kort 14 is er slechts een studie naar de cardiale nucleaire proteoom, en zij van mening was de hele kern, in plaats van het verkennen van het proteoom op het niveau van de nucleaire sub compartimenten 15. In groot deel is dit gebrek aan werk te wijten aan de moeilijkheid van het isoleren van cardiale kernen. Cardiac kernen optreden binnen een starre en dichte actine-myosine inrichting waaraan zij verbonden via meerdere verlengingen van het endoplasmatisch reticulum, voorzover myocyte contractie verandert bovendien hun algemene vorm. 16 cardiomyocyten zijn 40% mitochondria vol 17 die necessitates verrijking van de kern naast de andere organellen. Hier beschrijven we een protocol voor cardiale nucleaire verrijking en verdere fractionering in biologisch relevante compartimenten. Verder hebben we detail werkwijzen voor label-free kwantitatieve massaspectrometrische dissectie van deze fracties-technieken vatbaar voor in vivo experimenten in verschillende diermodellen en orgaansystemen waar metabolische labeling niet haalbaar.

Figuur 4 benadrukt het nut van deze vorm van relatieve kwantificatie. Getoond in het linker paneel zijn de individuele mono-isotopische peptide pieken (bedekt met verschillende muizen), die zijn aangewezen als behorend tot het eiwit HMGB1 (geïdentificeerd via database). Elke piek in wezen een ionenchromatograaf geëxtraheerd voor de gegeven peptide afkomstig is van een andere muis. De groepen vertegenwoordigen drie verschillende fysiologische toestanden: basale, cardiale hypertrofie en hartfalen, met d...

Watch this Scientific Journal Video about Quantitative Analysis of Chromatin Proteomes in Disease at JoVE.com

Quantitative Analysis of Chromatin Proteomes in Disease

Advances in massaspectrometrie toestemming hebben high throughput analyse van eiwitexpressie en modificatie in een groot aantal weefsels. Gecombineerd met subcellulaire fractionering en ziektemodellen, kwantitatieve massaspectrometrie en bioinformatica nieuwe eigenschappen kunnen onthullen in biologische systemen. De werkwijze hierin beschreven analyseert chromatine-geassocieerde eiwitten in de vaststelling van hart-en vaatziekten en is gemakkelijk toepasbaar op andere<em&gt; In vivo</em&gt; Modellen van menselijke ziekten.

Kwantitatieve analyse van chromatine proteomen in de ziekte van

In the nucleus reside the proteomes whose functions are most intimately linked with gene regulation. Adult mammalian cardiomyocyte nuclei are unique due ...

Research

JoVE Journal

Medicine

Quantitative Analysis of Cancer Metastasis using an Avian Embryo Model

Kwantitatieve analyse van kanker metastase met behulp van een Avian Embryo Model

During metastasis cancer cells disseminate from the primary tumor, invade into surrounding tissues, and spread to distant organs. Metastasis is a complex ...

A Simple Method of Mouse Lung Intubation

Een eenvoudige methode van de Muis van Lung intubatie

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

Quantitative Analysis and Characterization of Atherosclerotic Lesions in the Murine Aortic Sinus

Atherosclerosis is a disease of the large arteries and a major underlying cause of myocardial infarction and stroke. Several different mouse models have ...

Quantitative Magnetic Resonance Imaging of Skeletal Muscle Disease

Quantitative Magnetic Resonance Imaging van de skeletspieren Disease

Quantitative magnetic resonance imaging (qMRI) describes the development and use of MRI to quantify physical, chemical, and/or biological properties of ...

Quantitative Mass Spectrometric Profiling of Cancer-cell Proteomes Derived From Liquid and Solid Tumors

Kwantitatieve Massaspectrometrische Profilering van Kanker-cel proteoom Afgeleid Van vloeibare en vaste tumoren

In-depth analyses of cancer cell proteomes are needed to elucidate oncogenic pathomechanisms, as well as to identify potential drug targets and diagnostic ...

Quantitative 3D In Silico Modeling (q3DISM) of Cerebral Amyloid-beta Phagocytosis in Rodent Models of Alzheimer's Disease

Quantitative 3D In Silico Modeling q3DISM of Cerebral Amyloid-beta Phagocytosis in Rodent Models of Alzheimer's Disease

kwantitatieve 3D<em&gt; In Silico</em&gt; Modeling (q3DISM) van cerebrale amyloid-beta Phagocytosis in diermodellen van de ziekte van Alzheimer

Neuroinflammation is now recognized as a major etiological factor in neurodegenerative disease. Mononuclear phagocytes are innate immune cells responsible ...

Application of Granger Causality Analysis of the Directed Functional Connection in Alzheimer's Disease and Mild Cognitive Impairment

Toepassing van Granger causaliteit analyse van de gestuurde functionele verbinding in de ziekte van Alzheimer en Mild cognitief stoornis

Impaired functional connectivity in the Default Mode Network (DMN) may be involved in the progression of Alzheimer&#39;s Disease (AD). The Posterior ...

In Vivo Morphometric Analysis of Human Cranial Nerves Using Magnetic Resonance Imaging in Meni&#232;re's Disease Ears and Normal Hearing Ears

In Vivo Morphometric Analysis of Human Cranial Nerves Using Magnetic Resonance Imaging in MeniÃƒÆ’Ã†'Ãƒâ€šÃ‚Â¨re's Disease Ears and Normal Hearing Ears

In Vivo Morfometrische analyse van menselijke craniale zenuwen met behulp van magnetische resonantie beeldvorming in Menière de ziekte oren en normaal gehoor oren

Analysis of neural structures in Meni&#232;re's Disease (MD) is of importance, since a loss of such structures has previously been proposed for this ...

Whole-brain Segmentation and Change-point Analysis of Anatomical Brain MRI&#8212;Application in Premanifest Huntington's Disease

Whole-brain Segmentation and Change-point Analysis of Anatomical Brain MRIÃƒÆ’Ã‚Â¢ÃƒÂ¢Ã¢â‚¬Å¡Ã‚Â¬ÃƒÂ¢Ã¢â€šÂ¬Ã‚ÂApplication in Premanifest Huntington's Disease

Geheel-hersenen segmentatie en verandering-punt analyse van anatomische hersenen MRI — toepassing in Premanifest de ziekte van Huntington

Recent advances in MRI offer a variety of useful markers to identify neurodegenerative diseases. In Huntington&#39;s disease (HD), regional brain atrophy ...

Quantitative Micro-CT Analysis of Aortopathy in a Mouse Model of &#946;-aminopropionitrile-induced Aortic Aneurysm and Dissection

Quantitative Micro-CT Analysis of Aortopathy in a Mouse Model of ÃƒÆ’Ã…Â½Ãƒâ€šÃ‚Â²-aminopropionitrile-induced Aortic Aneurysm and Dissection

Kwantitatieve analyse van de Micro-CT van Aortopathy in een muismodel van β-aminopropionitrile-geïnduceerde aorta aneurysma en dissectie

Aortic aneurysm and dissection is associated with significant morbidity and mortality in the population and can be highly lethal. While animal models of ...