SWI/SNF and ISW2 have several basic catalytic activities in common such as their DNA translocase domains binding to the same location in nucleosomes and moving nucleosomes to different positions on DNA. But they also have important differences in nucleosome remodeling, such as whether they can conservatively move nucleosomes on DNA without altering the canonical structure (ISW2) or cause disruptions of histone–DNA interactions and even disassemble nucleosomes (SWI/SNF). While these differences may be orchestrated by the auxiliary subunits or domains of these complexes, we have found that even the interactions of the ATPase domains are distinct for these two complexes. Although the ATPase domains of SWI/SNF and ISW2 are highly similar at the level of their catalytic cores (motifs I–VI), they function quite differently making it seemingly impossible for one ATPase domain to replace another without serious problems. A fundamental difference is the interactions of the ATPase domain of ISW2 and Snf2 with nucleosomal DNA. The ATPase domain of ISW2 contacts the surface of nucleosomal DNA facing away from histone octamer, but SWI/SNF likely binds between nucleosomal DNA and the histone octamer. In this situation, SWI/SNF would have a strategic advantage for disrupting histone–DNA contacts with the ATPase domain positioned as a wedge that when translocating along DNA could readily break additional histone–DNA contacts.
The DNA footprint of ISW2-nucleosome shows discrete binding of ISW2 to the exposed side of one helical turn of nucleosomal DNA consistent with the ATPase domain binding to the outside surface of nucleosomal DNA (12
). The interactions of SWI/SNF with nucleosomal DNA by DNA footprinting are more extensive than that indicated by DNA crosslinking. Snf2 is primarily crosslinked to ~16
bp of nucleosomal DNA, but SWI/SNF protects a region of ~50
bp of nucleosomal DNA. This discrepancy may be due to the limited reactivity of the aryl azide and could require a more reactive crosslinker as shown previously (29
). Binding of SWI/SNF between the DNA gyre and histone octamer would require movement of the DNA and/or of the histone octamer to make space for Snf2. There is no indication as of yet that the DNA is significantly pulled off the histone octamer thereby losing its rotational phasing on the nucleosome to provide this additional flexibility, but further studies will be necessary to find how this might occur.
The DNA crosslinking experiments could be misleading if modification of the DNA altered the rotational phasing of nucleosomes. We suspect this is unlikely because the rotational phasing of nucleosomes is not readily changed, especially of strong nucleosome positioning sequences like 601. Evidence for rotational phasing likely not being altered by DNA modification comes from experiments with polyamides that bind to nucleosomal DNA. Polyamides bind to the minor groove of nucleosomes and can distort the DNA helix, but have been observed to not alter the protein–DNA interactions inside the nucleosome (38–40
). While binding of polyamides to SHL6 changed the rotational phasing at SHL-6, these changes were found to be due to crystallization conditions and as observed by DNase I footprinting likely do not reflect the state in solution. The attachment of an aryl azide to phosphate is a significantly smaller and less obtrusive to nucleosomal DNA and less likely to change the rotational phasing of nucleosomes.
Another concern for the DNA crosslinking experiments is the differences in nucleosomes with one being end positioned nucleosomes and the other more central positioned nucleosomes for crosslinking to SWI/SNF and ISW2. Extranucleosomal DNA seems to influence SWI/SNF as seen by SWI/SNF moving nucleosomes to preferred sites away from nucleosome positioning sequences (41–43
), but is not likely to change the way in which SWI/SNF engages nucleosomes in the absence of ATP. Unlike ISW2, SWI/SNF does not require extranucleosomal DNA in order to remodel nucleosomes and extranucleosomal DNA cannot be used to uniquely position SWI/SNF onto nucleosomes (15
). The key in these experiments is that the ATPase domains of Snf2 or Isw2 are shown to engage the same region of the nucleosome under these different conditions and thus provides us the opportunity to compare their binding under structurally similar conditions although different conditions have been used to achieve this purpose.
The functional consequence of the ATPase domain being bound on the outside of the nucleosome with no apparent change in histone–DNA interactions or wedged between DNA and histone octamer could affect the way which nucleosomes are moved on DNA (). As shown previously, after addition of ATP, SWI/SNF quickly disrupts histone–DNA contacts 54
bp from the dyad axis and they remain broken until the DNA has been moved 52
). These observations are consistent with SWI/SNF wedging between the DNA and octamer and upon translocation causing large scale disruptions of histone–DNA interactions. This same action could also cause DNA loops of significant size to form on the nucleosome surface. Extensive characterization of SWI/SNF remodeling intermediates by restriction enzyme accessibility has provided evidence for the formation of DNA loops during remodeling (19
). The formation of DNA loops large enough to be detected by restriction enzyme cutting is a property reserved for SWI/SNF and is not observed for ISWI remodelers. A concern raised about the DNA looping model for SWI/SNF and RSC remodeling has been the step size of the DNA translocase being so much smaller than the loop size being created (21
). Finding that the DNA translocase starts out wedge between the histone octamer and nucleosomal DNA accounts for how short movements on nucleosomal DNA could have longer range effects on histone–DNA interactions. A natural outcome of creating these large scale disruptions of histone–DNA interactions followed by repositioning the DNA back on to the histone octamer surface is that some differences in the path DNA are likely to occur. Distortions of this kind have been observed for SWI/SNF remodeling by high resolution mapping of specific histone–DNA contacts before and after remodeling to examine the spacing of DNA on nucleosomes (16
Figure 5. Model for the two modes of SWI/SNF and ISW2 remodeling. The nucleosome is shown with DNA in black and the ATPase domains of SWI/SNF and ISW2 as a dark grey spheres. SWI/SNF remodeling with the ATPase domain intercalated between the DNA gyre and histone (more ...)
ISW2 remodels nucleosomes in a very different manner that correlates well to its ATPase domain binding to the exposed side of nucleosomal DNA. Shortly after the addition of ATP, ISW2 moves DNA short distances of only 9–11
bp on the histone octamer without ever massively or persistently disrupting histone–DNA contacts like SWI/SNF (24
). The formation of DNA loops during ISW2 remodeling is also not detected by restriction enzyme cleavage. Likely due to the small and progressive changes in histone–DNA interactions that occur as part of ISW2 remodeling the pathway of DNA around the histone octamer is also well conserved after remodeling (18
). Translocation of the ATPase domain on the exposed surface as expected tends to move nucleosomes in short increments without significantly disrupting histone–DNA interactions. There may be a connection with the ATPase domain being lodge between the histone octamer and DNA that causes the remodeler to be more prone to disrupting histone–DNA interactions and unraveling DNA from the histone octamer such as for SWI/SNF. When the ATPase domain is instead bound to the outside surface of nucleosomal DNA it will likely move nucleosomes in such a way as to not disturb the canonical structure such as seen for ISW2.