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Elucidation of the functional role of the Q- and I-motif in the human chromatin-remodeling enzyme BRG1

The role of the Q-motif in BRG1 modeling
 
: Hoffmeister, Helen; Fuchs, Andreas; Strobl, Laura; Sprenger, Frank; Gröbner-Ferreira, Regina; Michaelis, Stefanie; Hoffmann, Petra; Nazet, Julian; Merkl, Rainer; Längst, Gernot

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The Journal of biological chemistry 294 (2019), Nr.9, S.3294-3310
ISSN: 0021-9258
ISSN: 1083-351X
Deutsche Forschungsgemeinschaft DFG
SFB 960;
Die Bildung von Ribosomen: Grundlagen der RNP-Biogenese und Kontrolle ihrer Funktion
Englisch
Zeitschriftenaufsatz
Fraunhofer EMFT ()
chromatin remodeling; tumor suppressor gene; ATP; nucleosome; cell proliferation; BRG1; cell cycle; ATPase; SF2 helicase family; ATPase domain

Abstract
The Snf2 proteins, comprising 53 different enzymes in humans, belong to the SF2 family. Many Snf2 enzymes possess chromatin remodeling activity, requiring a functional ATPase domain consisting of conserved motifs named Q and I-VII. These motifs form two recA-like domains, creating an ATP-binding pocket. Little is known about the function of the conserved motifs in chromatin remodeling enzymes. Here, we characterized the function of the Q- and I (Walker I)-motif in hBRG1 (SMARCA4). The motifs are in close proximity to the bound ATP, suggesting a role in nucleotide binding and/or hydrolysis. Unexpectedly, when substituting the conserved residues Q758 (Q-motif) or K785 (I-motif) of both motifs, all variants still bound ATP and exhibited basal ATPase activity similar to that of wild type BRG1 (wtBRG1). However, all mutants lost the nucleosome-dependent stimulation of the ATPase domain. Their chromatin remodeling rates were impaired accordingly, but nucleosome binding was retained and still comparable to that of wtBRG1. Interestingly, a cancer-relevant substitution, L754F (Q-motif) displayed defects similar to the Q-758-variant(s), arguing for a comparable loss of function. Since we excluded a mutual interference of ATP and nucleosome binding, we postulate that both motifs stimulate the ATPase and chromatin remodeling activities upon binding of BRG1 to nucleosomes, probably via allosteric mechanisms. Furthermore, mutations of both motifs similarly affect the enzymatic functionality of BRG1 in vitro and in living cells. Of note, in BRG1-deficient H1299 cells, exogenously expressed wtBRG1, but not BRG1 Q758A and BRG1 K785R, exhibited a tumor suppressor like function.

: http://publica.fraunhofer.de/dokumente/N-531922.html