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Genes to Cells (2009) 14, 1271-1330. doi:10.1111/j.1365-2443.2009.01350.x
© 2009 Blackwell Publishing or its licensors

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Global analysis of mutual interaction surfaces of nucleosomes with comprehensive point mutants

Makoto Sakamoto1{dagger}, Shuhei Noguchi1{dagger}, Satoshi Kawashima2{dagger}, Yusuke Okada2, Takemi Enomoto2, Masayuki Seki2 and Masami Horikoshi1,*

1 Laboratory of Developmental Biology, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan
2 Molecular Cell Biology Laboratory, Graduate School of Pharmaceutical Sciences, Tohoku University, 6-3 Aoba, Aramaki, Aoba-ku, Sendai, Miyagi 980-8578, Japan

The surfaces of core histones in nucleosome are exposed as required for factor recognition, or buried for histone–DNA and histone–histone interactions. To understand the mechanisms by which nucleosome structure and function are coordinately altered in DNA-mediated reactions, it is essential to define the roles of both exposed and buried residues and their functional relationships. For this purpose, we developed GLASP (GLobal Analysis of Surfaces by Point mutation) and GLAMP (GLobal Analysis of Mutual interaction surfaces of multi-subunit protein complex by Point mutation) strategies, both of which are comprehensive analyses by point mutagenesis of exposed and buried residues in nucleosome, respectively. Four distinct DNA-mediated reactions evaluated by Ty suppression (the Spt phenotype), and sensitivities to 6-azauracil (6AU), hydroxyurea (HU), and methyl methanesulfonate (MMS), require common and different GLAMP residues. Mutated GLAMP residues at the interface between histones H2A and H2B mainly affect the Spt phenotype but not HU and MMS sensitivities. Interestingly, among the mutated GLAMP residues surrounding the histone H3–H3' interface, some equally affect the Spt phenotype, and HU and MMS sensitivities, whereas others differentially affect the Spt phenotype, and HU and MMS sensitivities. Based on these and other results, the functional relationships among chromatin factors and GLASP and GLAMP residues provide insights into nucleosome disassembly/assembly processes in DNA-mediated reactions.


*Correspondence: horikosh{at}iam.u-tokyo.ac.jp

{dagger}These authors contributed equally to this work.

Communicated by: Hiroshi Handa







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