Chromatin structure and induction-dependent conformational changes of human interferon-β genes in a mouse host cell

Authors


Correspondence to J. Bode, Abteilung Genetik, GBF, Gesellschaft für Biotechnologische Forschung mbH, Mascheroder Weg 1, D-3300 Braunschweig-Stöckheim, Federal Republic of Germany

Abstract

Multiple copies of a human interferon-β gene introduced into a mouse host cell line can be activated by induction with double-stranded RNA. Several induction-dependent changes of the chromatin structure could be traced by mapping techniques using four different agents [DNase I, micrococcus nuclease, bromoacetaldehyde and methidiumpropyl-EDTA · iron(II)]. Our data show that all copies of the interferon gene have adopted a very similar conformation in the host cell and respond to the inducing stimulus in a highly synchronous fashion. Detailed induction-specific changes were observed best with the chemical reagents which disclose a specific hypersensitive site within a sequence that has been shown to be required for the induction process (around position –80) and three other regions which, in addition to the transcribed region itself, gain single-strand character by an auxiliary process which can be mimicked by the addition of butyrate to the medium and may therefore involve histone hyperacetylation. Six discrete ‘phased’ nucleosomes are present upstream from the gene and are modulated by induction. At least four nucleosomes are located downstream. The interferon genes are largely protected from micrococcus nuclease in the inactive state. Gene activation increases access to micrococcus nuclease and DNase I indicating gross conformational changes on a higher level of chromatin structure.

Abbreviations
bp

base pairs

kb

103 bases

MPE

methidiumpropyl-EDTA · iron(II)

PhMeSO2F

phenylmethylsulfonyl fluoride

SDS

sodium dodecyl sulfate

Enzymes
 

(IUB Recommendations, 1984). DNase I (EC 3.1.21.1)

 

nuclease S1 (EC 3.1.30.1)

 

micrococcal nuclease (EC 3.1.31.1)

 

proteinase K (EC 3.4.21.14)

 

RNase A (EC 3.1.27.5)

 

restriction endonucleases

 

BglII, EcoRI, HindIII, HincII, HpaI, NcoI, PstI (EC 3.1.21.4)

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