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Towards elucidating the stability, dynamics and architecture of the nucleosome remodeling and deacetylase complex by using quantitative interaction proteomics
Author(s) -
Kloet Susan L.,
Baymaz H. Irem,
Makowski Matthew,
Groenewold Vincent,
Jansen Pascal W. T. C.,
Berendsen Madeleine,
Niazi Hassin,
Kops Geert J.,
Vermeulen Michiel
Publication year - 2015
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.12972
Subject(s) - stable isotope labeling by amino acids in cell culture , nucleosome , protein subunit , histone , biology , chromatin remodeling , chromatin , histone deacetylase , chemistry , microbiology and biotechnology , biochemistry , biophysics , proteomics , dna , gene
The nucleosome remodeling and deacetylase (Nu RD ) complex is an evolutionarily conserved chromatin‐associated protein complex. Although the subunit composition of the mammalian complex is fairly well characterized, less is known about the stability and dynamics of these interactions. Furthermore, detailed information regarding protein–protein interaction surfaces within the complex is still largely lacking. Here, we show that the Nu RD complex interacts with a number of substoichiometric zinc finger‐containing proteins. Some of these interactions are salt‐sensitive ( ZNF 512B and SALL 4), whereas others ( ZMYND 8) are not. The stoichiometry of the core subunits is not affected by high salt concentrations, indicating that the core complex is stabilized by hydrophobic interactions. Interestingly, the RBBP 4 and RBBP 7 proteins are sensitive to high nonionic detergent concentrations during affinity purification. In a subunit exchange assay with stable isotope labeling by amino acids in cell culture ( SILAC )‐treated nuclear extracts, RBBP 4 and RBBP 7 were identified as dynamic core subunits of the Nu RD complex, consistent with their proposed role as histone chaperones. Finally, using cross‐linking MS , we have uncovered novel features of Nu RD molecular architecture that complement our affinity purification‐ MS / MS data. Altogether, these findings extend our understanding of MBD 3–Nu RD structure and stability. Structured digital abstractMBD3 physically interacts with ZNF512B , HDAC1 , ZMYND8 , GATAD2B , SALL4 , GATAD2A , ZNF592 , MTA3 , ZNF687 , CDK2AP1 , CHD3 , ZNF532 , HDAC2 , MTA2 , CHD4 , MTA1 , KPNA2 , CHD5 , RBBP4 and RBBP7 by pull down ( View interaction ) CDK2AP1 physically interacts with MBD3 , MTA3 , HDAC2 , GATAD2A , CHD4 , CDK2AP1 , MTA2 , HDAC1 , MTA1 , CHD3 , GATAD2B , MBD2 , RBBP4 and RBBP7 by pull down ( View interaction ) MBD3 physically interacts with MTA2 , MTA3 , RBBP4 , RBBP7 , HDAC2 , HDAC1 , CHD4 , CHD3 and MTA1 by cross-linking study ( View interaction )