Etudes sur le mécanisme de remodelage des nucléosomes par ...

tel-00413908, version 1 - 7 Sep 2009
I.5.1.1 Substrate binding
In order to remodel the nucleosomes the chromatin remodelers must recognize their
substrates. It is expected from an enzyme that brings about changes in chromatin to interact
with DNA. Initial methods to isolate human SWI/SNF included DNA affinity columns
indicating towards a nonspecific binding of these complexes to DNA (Kwon et al., 1994). The
nucleosome binding activity was evident from initial gel shift experiments with yeast
SWI/SNF complex (Côté et al., 1994). Later, the binding properties of both SWI/SNF and
ISWI group of remodelers have been established. SWI/SNF remodelers display an affinity of
~10 -8 M -1 for DNA substrates in an ATP dependent manner (Quinn et al., 1996; Lorch et al.,
1998). No difference in binding affinity between DNA and nucleosomes was observed.
However, the binding affinity to nucleosomes increases more than three fold in presence of
ATP (Lorch et al., 1998). Similarly, RSC remodeling complex does not show any preference
for the presence of linker DNA for binding. For ISWI group of remodelers the binding
preferences are slightly different. ISWI can bind DNA but with a lower affinity than
SWI/SNF group of remodelers (Whitehouse et al., 2003). Moreover, the presence of linker
DNA increases the binding affinity towards the nucleosomes (Brehm et al., 2000). It is known
that SWI/SNF exhibits a high affinity for four way junction (4WJ) DNA. This property is
similar to as shown by HMG-box domain proteins (Quinn et al., 1996). It is noteworthy that
this structure is very similar to the entry exit site nucleosomal DNA. Therefore, it was
proposed that SWI/SNF and related complexes may bind the entry exit segment of
nucleosomal DNA.
Further details about the nucleosome binding of remodelers have been obtained using
structural studies using cross linking (Sengupta et al., 2001) and Electron microscopy
methods. It has been shown that ISWI contacts three distinct regions within the nucleosomal
DNA (i)~10bp of nucleosomal DNA at super helical location 2 (SHL2); (ii) 10 bp region near
the entry exit site of DNA and (iii) linker DNA (Kagalwala et al., 2004). SWI/SNF makes
contact with ~60 bp of nucleosomal DNA from entry site of DNA to SHL2 (Dechassa et al.,
2008). Similarly, RSC has been shown to interact with DNA near the SHL2, however, the
interaction data was based on DNaseI footprinting experiment and needs to be confirmed by
definitive cross-linking studies (Saha et al., 2006).
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