1. Volume 62, Issue 3, Pages 453-461 (May 2016)
Regulation of DNA Translocation Efficiency within the Chromatin Remodeler RSC/Sth1 Potentiates Nucleosome Sliding and Ejection 
Cedric R. Clapier, Margaret M. Kasten, Timothy J. Parnell, Ramya Viswanathan, Heather Szerlong, George Sirinakis, Yongli Zhang, Bradley R. Cairns 
Molecular Cell 
Volume 62, Issue 3, Pages (May 2016)
DOI: /j.molcel
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2. Molecular Cell 2016 62, 453-461DOI: (10.1016/j.molcel.2016.03.032)
Copyright © 2016 Elsevier Inc. Terms and Conditions

3. Figure 1 Actin-Related Proteins Promote Nucleosome Ejection by RSC
(A) RSC, with core motor subunits colored and accessory subunits in gray. Below, the two alternative outcomes of nucleosome remodeling by RSC. At right, scheme of RSC core motor subunits and their interactions (note: Rtt102-biases ARP conformations, see main text) and conserved Sth1 regulatory domains, with outset (below) depicting the mutations derived and investigated.
(B) Comparative sliding and ejection of mononucleosomes by RSC derivatives. Enzyme:substrate molar ratio is 1:2. ARP module re-addition (right panel) involves a 2-fold molar ratio over RSC ΔARPs. Bracket∗; (here, and upper region of native gels) other remodeling intermediates (e.g RSC/Sth1-nucleosome complexes). Here, and in subsequent figures, a representative gel from multiple experiments is shown.
(C) Principle of the nucleosome array ejection assay, with supercoiled plasmid (topoisomer) distribution revealed by 2D gel. Lk: Linking number, N: Nicked, L: Linear.
(D) Comparative nucleosome array ejection by RSC derivatives, (1:2 RSC:Nuc molar ratio; 90 min). Raw gel provided in Figure S10A.
See also Figures S1, S2, and S4.
Molecular Cell  , DOI: ( /j.molcel )
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4. Figure 2
ARPs Promote Coupling, while the Post-HSA Domain Regulates ATPase Activity
(A) Dominant lethality (red) conferred by expression of particular STH1Post-HSA alleles or by combining two mutations that separately confer arpΔ suppression (green).
(B) Comparative “Tet-tethered” DNA-translocation activity measured by accumulation of plasmid supercoiled (SC) topoisomers (see Figure S3A), with plasmid-stimulated ATPase activity (Av, blue text; represented as mean ± SEM) and coupling value (Cv, black text). Background color reflects classification into “Types” (see Figures 4 and S8; Discussion). At left, S: Sth1 alone; SA: Sth1+ARPs; SAR: Sth1+ARPs+Rtt102. SC: highly supercoiled topoisomers, R: Relaxed plasmid, M: Marker.
(C) Comparative sliding and ejection of mononucleosomes (S, SA, SAR formats). Enzyme:substrate molar ratio is 1:2.
(D) Single molecule measurements (optical tweezers) reveal enhanced velocity of DNA translocation with L392V or L392P mutation (SA format).
(E) Comparative nucleosome array ejection. ∗WT samples derived from the same experiment, run on a separate gel. Raw gels provided in Figure S10B.
(F) Impact on nucleosome positioning in vivo following expression of Sth1 WT or L392P, displaying nucleosome midpoints for promoters (heatmap, top and middle panels) or mean nucleosomal profile (bottom panel); data are from one typical cluster (cluster#3; see Figure S6 for all clusters).
See also Figures S1–S6 and S10.
Molecular Cell  , DOI: ( /j.molcel )
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5. Figure 3
The Protrusion 1 Domain Integrates ATP Hydrolysis and Coupling, and Synergizes with the Post-HSA
(A) The Protrusion 1 domain mediates and enhances coupling. Comparative DNA-translocation, plasmid-stimulated ATPase activity (Av, blue text) and coupling values (Cv, black text), as in Figure 2B. Background color reflects classification into “Types” (see Figure 4). SC: highly supercoiled topoisomers, R: Relaxed plasmid, M: Marker.
(B) Protrusion 1 suppressor mutations enhance nucleosome sliding. Comparative sliding and ejection of mononucleosomes (S format). Enzyme:substrate molar ratio is 1:2.
(C) Protrusion 1 suppressor mutations moderately enhance array ejection. ∗Samples from the same experiment, run on a different gel. Raw gels provided in Figure S10C.
(D–F) Combining Post-HSA and Protrusion 1 suppressor mutations increases DNA translocation (conditions and display as in Figure 3A, [D]) and confers strong ejection in mononucleosome (E) or array (F) assays. (S format; conditions as in [B].) Note: the L681F and control samples are the same in (C) and (F). Raw gels provided in Figure S10C.
See also Figure S7 for an additional mutant combination.
Molecular Cell  , DOI: ( /j.molcel )
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6. Figure 4
ARPs Promote Nucleosome Ejection by RSC—and the Integration of ATPase and Coupling Potentiates Nucleosome Sliding and Ejection
(A) ARPs enhance sliding (not depicted) and enable ejection in a nucleosome array format.
(B) A model for the integration of ATPase and coupling in remodeling outcomes. Summary of the impact of Sth1 regulatory domains (Post-HSA and Protrusion 1) and interacting proteins (ARPs and Rtt102) on DNA translocation and nucleosome remodeling (sliding and ejection). Different ATPase and coupling activities were grouped into four Input “Types.” A colored tachometer represents ATPase and Coupling activities/values, and the same color spectrum displays the impact on sliding and ejection. Red: none. Orange: low/weak. Yellow: moderate. Light Green: high/strong. Dark Green: very high. The black arcs represent the range of values displayed by the different Sth1 complexes (WT or mutants, +/− ARPs and Rtt102) within these Types. See Table S1 for all ATPase and coupling activities/values, and Figure S8 for examples of Input Types linked to the model.
See also Figures S8 and S9 and Table S1.
Molecular Cell  , DOI: ( /j.molcel )
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