全部 标题 作者
关键词 摘要

OALib Journal期刊
ISSN: 2333-9721
费用:99美元

查看量下载量

相关文章

更多...
PLOS ONE  2011 

Direct Identification of Insulator Components by Insertional Chromatin Immunoprecipitation

DOI: 10.1371/journal.pone.0026109

Full-Text   Cite this paper   Add to My Lib

Abstract:

Comprehensive understanding of mechanisms of epigenetic regulation requires identification of molecules bound to genomic regions of interest in vivo. However, non-biased methods to identify molecules bound to specific genomic loci in vivo are limited. Here, we applied insertional chromatin immunoprecipitation (iChIP) to direct identification of components of insulator complexes, which function as boundaries of chromatin domain. We found that the chicken β-globin HS4 (cHS4) insulator complex contains an RNA helicase protein, p68/DDX5; an RNA species, steroid receptor RNA activator 1; and a nuclear matrix protein, Matrin-3, in vivo. Binding of p68 and Matrin-3 to the cHS4 insulator core sequence was mediated by CCCTC-binding factor (CTCF). Thus, our results showed that it is feasible to directly identify proteins and RNA bound to a specific genomic region in vivo by using iChIP.

References

[1]  van Driel R, Fransz PF, Verschure PJ (2003) The eukaryotic genome: a system regulated at different hierarchical levels. J Cell Sci 116: 4067–4075.
[2]  Zhang XY, Horz W (1982) Analysis of highly purified satellite DNA containing chromatin from the mouse. Nucleic Acids Res 10: 1481–1494.
[3]  Workman JL, Langmore JP (1985) Nucleoprotein hybridization: a method for isolating specific genes as high molecular weight chromatin. Biochemstry 24: 7486–7497.
[4]  Boffa LC, Carpaneto EM, Allfrey VG (1995) Isolation of active genes containing CAG repeats by DNA strand invasion by a peptide nucleic acid. Proc Natl Acad Sci USA 92: 1901–1905.
[5]  Jaskinskas A, Hamkalo BA (1999) Purification and initial characterization of primate satellite chromatin. Chromosome Res 7: 341–354.
[6]  Griesenbeck J, Boeger H, Strattan JS, Kornburg RD (2003) Affinity purification of specific chromatin segments from chromosomal loci in yeast. Mol Cell Biol 23: 9275–9282.
[7]  Ghirlando R, Felsenfeld G (2008) Hydrodynamic studies on defined heterochromatin fragments support a 30-μm fiber having six nucleosomes per turn. J Mol Biol 376: 1417–1425.
[8]  Déjardin J, Kingston RE (2009) Purification of proteins associated with specific genomic Loci. Cell 136: 175–186.
[9]  Felsenfeld G, Burgess-Beusse B, Farrell C, Gaszner M, Ghirlando R, et al. (2004) Chromatin boundaries and chromatin domains. Cold Spring Harb Symp Quant Biol 69: 245–250.
[10]  Bushey AM, Dorman ER, Corces VG (2008) Chromatin insulators: regulatory mechanisms and epigenetic inheritance. Mol Cell 32: 1–9.
[11]  Bell AC, West AG, Felsenfeld G (1999) The protein CTCF is required for the enhancer blocking activity of vertebrate insulators. Cell 98: 387–396.
[12]  Hark AT, Schoenherr CJ, Katz DJ, Ingram RS, Levorse JM, et al. (2000) CTCF mediates methylation-sensitive enhancer-blocking activity at the H19/Igf2 locus. Nature 405: 486–489.
[13]  Gaszner M, Felsenfeld G (2006) Insulators: exploiting transcriptional and epigenetic mechanisms. Nat Rev Genet 7: 703–713.
[14]  Hoshino A, Fujii H (2009) Insertional chromatin immunoprecipitation: a method for isolating specific genomic regions. J Biosci Bioeng 108: 446–449.
[15]  Chung JH, Bell AC, Felsenfeld G (1997) Characterization of the chicken β-globin insulator. Proc Natl Acad Sci USA 94: 575–580.
[16]  Hanawa H, Yamamoto M, Zhao H, Shimada T, Persons DA (2009) Optimized lentiviral vector design improves titer and transgene expression of vectors containing the chicken β-globin locus HS4 insulator element. Mol Ther 17: 667–674.
[17]  Palacios R, Steinmetz M (1985) Il-3-dependent mouse clones that express B-220 surface antigen, contain Ig genes in germ-line configuration, and generate B lymphocytes in vivo. Cell 41: 727–734.
[18]  Fuller-Pace FV, Ali S (2008) The DEAD box RNA helicases p68 (Ddx5) and p72 (Ddx17): novel transcriptional co-regulators. Biochem Soc Trans 36: 609–612.
[19]  Belgrader P, Dey R, Berezney R (1991) Molecular cloning of matrin 3. A 125-kilodalton protein of the nuclear matrix contains an extensive acidic domain. J Biol Chem 266: 9893–9899.
[20]  Yusufzai TM, Tagami H, Nakatani Y, Felsenfeld G (2004) CTCF tethers an insulator to subnuclear sites, suggesting shared insulator mechanisms across species. Mol Cell 13: 291–298.
[21]  Lei EP, Corces VG (2006) RNA interference machinery influences the nuclear organization of a chromatin insulator. Nat Genet 38: 936–941.
[22]  Yao H, Brick K, Evrard Y, Xiao T, Camerini-Otero RD, et al. (2010) Mediation of CTCF transcriptional insulation by DEAD-box RNA-binding protein p68 and steroid receptor RNA activator SRA. Genes Dev 24: 2543–2555.
[23]  Suzuki T, Fujisawa J, Toita M, Yoshida M (1993) The trans-activator tax of human T-cell leukemia virus type 1 (HTLV-1) interacts with cAMP-responsive element (CRE) binding and CRE modulator proteins that bind to the 21-base-pair enhancer of HTLV-1. Proc Natl Acad Sci USA 90: 610–614.
[24]  Gerasimova TI, Byrd K, Corces VG (2000) A chromatin insulator determines the nuclear localization of DNA. Mol Cell 6: 1025–1035.
[25]  Ishii K, Arib G, Lin C, Van Houwe G, Laemmli UK (2002) Chromatin boundaries in budding yeast: the nuclear pore connection. Cell 109: 551–562.
[26]  Yusufzai TM, Felsenfeld G (2004) The 5′-HS4 chicken β-globin insulator is a CTCF-dependent nuclear matrix-associated element. Proc Natl Acad Sci USA 101: 8620–8624.
[27]  Kurokawa R, Rosenfeld MG, Glass CK (2009) Transcriptional regulation through noncoding RNAs and epigenetic modifications. RNA Biol 6: 233–236.
[28]  Hoshino A, Matsumura S, Kondo K, Hirst JA, Fujii H (2004) Inducible translocation trap: a system to detect translocation of signaling molecules induced by extracellular stimuli. Mol Cell 15: 153–159.
[29]  Yahata K, Maeshima K, Sone T, Ando T, Okabe M, et al. (2007) cHS4 insulator-mediated alleviation of promoter interference during cell-based expression of tandemly associated transgenes. J Mol Biol 374: 580–590.
[30]  Hoshino A, Fujii H (2007) Redundant promoter elements mediate IL-3-induced expression of a novel cytokine-inducible gene, cyclon. FEBS Lett 581: 975–980.
[31]  Fujita T, Ryser S, Tortola S, Piuz I, Schlegel W (2007) Gene-specific recruitment of positive and negative elongation factors during stimulated transcription of the MKP-1 gene in neuroendocrine cells. Nucleic Acids Res 35: 1007–1017.

Full-Text

Contact Us

service@oalib.com

QQ:3279437679

WhatsApp +8615387084133