Abstract
The highly dynamic nucleoprotein structure of eukaryotic genome is organized in an ordered fashion, the unit of which is the nucleosome. The nucleosome is composed of core histones and DNA of variable size wrapped around it. Apart from the histone proteins, several nonhistone proteins also interact with the complex consisting of the DNA, the core and linker histones conferring highly regulated fluidity on the chromatin and permitting fine tuning of its functions. The nonhistone proteins are multifunctional and accentuate diverse cellular outcomes. In spite of the technical challenges, the architectural role of the nonhistone proteins altering the topology of the chromatin has been studied extensively. To appreciate the significance of the chromatin for genome function, it is essential to examine the role of the nonhistone proteins in different physiological conditions. Here, taking the example of a highly abundant chromatin protein, PC4 (Positive coactivator 4), we describe strategies for the identification of the chromatin-associated proteins and their structural and functional characterization.
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References
Wolffe AP, Khochbin S, Dimitrov S (1997) What do linker histones do in chromatin? Bioessays 19:249–255
Bannister AJ, Zegerman P, Partridge JF, Miska EA, Thomas JO, Allshire RC, Kouzarides T (2001) Selective recognition of methylated lysine 9 on histone H3 by the HP1 chromo domain. Nature 410:120–124
Lachner M, O’Carroll D, Rea S, Mechtler K, Jenuwein T (2001) Methylation of histone H3 lysine 9 creates a binding site for HP1 proteins. Nature 410:116–120
Grigoryev SA, Bednar J, Woodcock CL (1999) MENT, a heterochromatin protein that mediates higher order chromatin folding, is a new serpin family member. J Biol Chem 274:5626–5636
Springhetti EM, Istomina NE, Whisstock JC, Nikitina T, Woodcock CL, Grigoryev SA (2003) Role of the M-loop and reactive center loop domains in the folding and bridging of nucleosome arrays by MENT. J Biol Chem 278:43384–43393
Waldmann T, Baack M, Richter N, Gruss C (2003) Structure-specific binding of the proto-oncogene protein DEK to DNA. Nucleic Acids Res 31:7003–7010
Alexiadis V, Waldmann T, Andersen J, Mann M, Knippers R, Gruss C (2000) The protein encoded by the proto-oncogene DEK changes the topology of chromatin and reduces the efficiency of DNA replication in a chromatin-specific manner. Genes Dev 14:1308–1312
Ge H, Roeder RG (1994) Purification, cloning, and characterization of a human coactivator, PC4, that mediates transcriptional activation of class II genes. Cell 78:513–523
Kretzschmar M, Kaiser K, Lottspeich F, Meisterernst M (1994) A novel mediator of class II gene transcription with homology to viral immediate-early transcriptional regulators. Cell 78:525–534
Pan ZQ, Ge H, Amin AA, Hurwitz J (1996) Transcription-positive cofactor 4 forms complexes with HSSB (RPA) on single-stranded DNA and influences HSSB-dependent enzymatic synthesis of simian virus 40 DNA. J Biol Chem 271:22111–22116
Wang JY, Sarker AH, Cooper PK, Volkert MR (2004) The single-strand DNA binding activity of human PC4 prevents mutagenesis and killing by oxidative DNA damage. Mol Cell Biol 24:6084–6093
Micheli L, Leonardi L, Conti F, Buanne P, Canu N, Caruso M, Tirone F (2005) PC4 coactivates MyoD by relieving the histone deacetylase 4-mediated inhibition of myocyte enhancer factor 2C. Mol Cell Biol 25:2242–2259
Werten S, Stelzer G, Goppelt A, Langen FM, Gros P, Timmers HT, Van der Vliet PC, Meisterernst M (1998) Interaction of PC4 with melted DNA inhibits transcription. EMBO J 17:5103–5111
Brandsen J, Werten S, van der Vliet PC, Meisterernst M, Kroon J, Gros P (1997) C-terminal domain of transcription cofactor PC4 reveals dimeric ssDNA binding site. Nat Struct Biol 4:900–903
Das C, Hizume K, Batta K, Kumar BR, Gadad SS, Ganguly S, Lorain S, Verreault A, Sadhale PP, Takeyasu K, Kundu TK (2006) Transcriptional coactivator PC4, a chromatin-associated protein, induces chromatin condensation. Mol Cell Biol 26:8303–8315
Banerjee S, Kumar BR, Kundu TK (2004) General transcriptional coactivator PC4 activates p53 function. Mol Cell Biol 24:2052–2062
Batta K, Yokokawa M, Takeyasu K, Kundu TK (2009) Human transcriptional coactivator PC4 stimulates DNA end joining and activates DSB repair activity. J Mol Biol 385:788–799
Das C, Gadad SS, Kundu TK (2010) Human positive coactivator 4 controls heterochromatinization and silencing of neural gene expression by interacting with REST/NRSF and CoREST. J Mol Biol 397:1–12
Diaz R, Stahl PD (1989) Digitonin permeabilization procedures for the study of endosome acidification and function. Methods Cell Biol 31:25–43
Kingston RE, Chen CA, Okayama H (2001) Calcium phosphate transfection. Curr Protoc Immunol Chapter 10, Unit 10.13
Acknowledgements
The study presented here is supported by Department of Biotechnology (DBT) and Jawaharlal Nehru Centre for Advanced Scientific Research, Government of India. SK is a CSIR (Council of Scientific and Industrial Research) Senior Research Fellow. TKK is a Sir J.C. Bose National Fellow. We acknowledge Prof. M. R. S. Rao, President, Jawaharlal Nehru Centre for Advanced Scientific Research, for his insightful suggestions in the study of PC4 as a chromatin protein.
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Kumari, S. et al. (2015). Identification and Characterization of Nonhistone Chromatin Proteins: Human Positive Coactivator 4 as a Candidate. In: Chellappan, S. (eds) Chromatin Protocols. Methods in Molecular Biology, vol 1288. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2474-5_14
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DOI: https://doi.org/10.1007/978-1-4939-2474-5_14
Publisher Name: Humana Press, New York, NY
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