Harpprecht, L.* ; Baldi, S.* ; Schauer, T.* ; Schmidt, A.* ; Bange, T.* ; Robles, M.S.* ; Kremmer, E. ; Imhof, A.* ; Becker, P.B.*
     
 
    
        
A Drosophila cell-free system that senses DNA breaks and triggers phosphorylation signalling.
    
    
        
    
    
        
        Nucleic Acids Res. 47, 7444-7459 (2019)
    
    
    
		
		
			
				Preblastoderm Drosophila embryo development is characterized by fast cycles of nuclear divisions. Extracts from these embryos can be used to reconstitute complex chromatin with high efficiency. We now discovered that this chromatin assembly system contains activities that recognize unprotected DNA ends and signal DNA damage through phosphorylation. DNA ends are initially bound by Ku and MRN complexes. Within minutes, the phosphorylation of H2A.V (homologous to gamma H2A.X) initiates from DNA breaks and spreads over tens of thousands DNA base pairs. The gamma H2A.V phosphorylation remains tightly associated with the damaged DNA and does not spread to undamaged DNA in the same reaction. This first observation of long-range gamma H2A.X spreading along damaged chromatin in an in vitro system provides a unique opportunity for mechanistic dissection. Upon further incubation, DNA ends are rendered single-stranded and bound by the RPA complex. Phosphoproteome analyses reveal damage-dependent phosphorylation of numerous DNA-end-associated proteins including Ku70, RPA2, CHRAC16, the exonuclease Rrp1 and the telomer capping complex. Phosphorylation of spindle assembly checkpoint components and of microtubule-associated proteins required for centrosome integrity suggests this cell-free system recapitulates processes involved in the regulated elimination of fatally damaged syncytial nuclei.
			
			
				
			
		 
		
			
				
					
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        Publikationstyp
        Artikel: Journalartikel
    
 
    
        Dokumenttyp
        Wissenschaftlicher Artikel
    
 
    
        Typ der Hochschulschrift
        
    
 
    
        Herausgeber
        
    
    
        Schlagwörter
        Double-strand Breaks; Chromatin Accessibility; Xenopus-laevis; Messenger-rna; Repair; Complex; Protein; Damage; Dynamics; Melanogaster
    
 
    
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        Sprache
        englisch
    
 
    
        Veröffentlichungsjahr
        2019
    
 
    
        Prepublished im Jahr 
        
    
 
    
        HGF-Berichtsjahr
        2019
    
 
    
    
        ISSN (print) / ISBN
        0305-1048
    
 
    
        e-ISSN
        1362-4962
    
 
    
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	    Band: 47,  
	    Heft: 14,  
	    Seiten: 7444-7459 
	    Artikelnummer: ,  
	    Supplement: ,  
	
    
 
  
        
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            Verlag
            Oxford University Press
        
 
        
            Verlagsort
            Great Clarendon St, Oxford Ox2 6dp, England
        
 
	
        
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        Begutachtungsstatus
        Peer reviewed
    
 
     
    
        POF Topic(s)
        30504 - Mechanisms of Genetic and Environmental Influences on Health and Disease
    
 
    
        Forschungsfeld(er)
        Immune Response and Infection
    
 
    
        PSP-Element(e)
        G-501793-001
    
 
    
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        Erfassungsdatum
        2019-09-24