Abstract
Conserved ATP-dependent chromatin remodelers establish and maintain genome-wide chromatin architectures of regulatory DNA during cellular lifespan, but the temporal interactions between remodelers and chromatin targets have been obscure. We performed live-cell single-molecule tracking for RSC, SWI/SNF, CHD1, ISW1, ISW2, and INO80 remodeling complexes in budding yeast and detected hyperkinetic behaviors for chromatin-bound molecules that frequently transition to the free state for all complexes. Chromatin-bound remodelers display notably higher diffusion than nucleosomal histones, and strikingly fast dissociation kinetics with 4-7 s mean residence times. These enhanced dynamics require ATP binding or hydrolysis by the catalytic ATPase, uncovering an additional function to its established role in nucleosome remodeling. Kinetic simulations show that multiple remodelers can repeatedly occupy the same promoter region on a timescale of minutes, implicating an unending 'tug-of-war' that controls a temporally shifting window of accessibility for the transcription initiation machinery.
Highlights
; Kubik et al, 2018; Yen et al, 2012)
Imaging chromatin remodeler diffusion by the fast-tracking mode in yeast shows that they bind to chromatin at substantial frequencies [Fbound: 35 - 66%], and with a notable population [21 - 30%] displaying intermediate D values resulting from transitions between bound and free states (Figure 1-3)
We speculate that the multiplicity of interaction motifs has a central role in the unusual diffusive behaviors shown by chromatin remodelers. 432 Importantly, the fast dissociation rates of remodelers are facilitated by ATP hydrolysis
Summary
403 Imaging chromatin remodeler diffusion by the fast-tracking mode in yeast shows that they bind to chromatin at substantial frequencies [Fbound: 35 - 66%], and with a notable population [21 - 30%] displaying intermediate D values resulting from transitions between bound and free states (Figure 1-3). Five tested ATPase-dead mutants (for ISW2, ISW1, CHD1) show two-fold or greater increase in their stable-binding residence times (Figure 5C-F), highlighting a new role of for ATP-utilization in coupling nucleosome remodeling to rapid enzyme dissociation from chromatin. This suggests that their mean residence times can reflect timescales for the diverse reactions performed by remodeling enzymes on chromatin in vivo. We suggest that there may be robust and dynamic competition between PIC components and mobilized NDR-flanking nucleosomes with chromatin exposure of key promoter elements such as the TATA box occurring for only a limited time window allowing proper assembly of downstream PIC components This temporally positioned +1 nucleosome would enable Pol II to scan and start transcription at the proper, canonical TSS. Our findings in live cells provide a temporal framework for further testing of proposed models and should facilitate development of in vitro single-molecule assays that allow direct observation of physical and functional interactions between transcription regulators, chromatin, and the transcription machinery
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