Binding of G-quadruplexes to the N-terminal Recognition Domain of the RNA Helicase Associated with AU-rich Element (RHAU)

Meier, Markus; Patel, Trushar R.; Booy, Evan P.; Marushchak, Oksana; Okun, Natalie; Deo, Soumya; Howard, Ryan; McEleney, Kevin; Harding, Stephen E.; Stetefeld, Jörg; McKenna, Sean Andrew

doi:10.1074/jbc.m113.512970

Cited by 53 publications

(97 citation statements)

References 49 publications

(170 reference statements)

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“…The most highly enriched protein in the G4DNA sample was the DHX36/RHAU/G4R1 RNA helicase, previously identified as the major source of G4DNA binding and unwinding activity in the cell (27)(28)(29). DHX36 is a multifunctional enzyme involved in transcription (30) and translation (31) and has been suggested to serve as a sensor for DNA in the cytoplasm (32).…”

Section: Resultsmentioning

confidence: 99%

Evidence That G-quadruplex DNA Accumulates in the Cytoplasm and Participates in Stress Granule Assembly in Response to Oxidative Stress

Byrd¹,

Zybailov

Maddukuri³

et al. 2016

Journal of Biological Chemistry

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Cells engage numerous signaling pathways in response to oxidative stress that together repair macromolecular damage or direct the cell toward apoptosis. As a result of DNA damage, mitochondrial DNA or nuclear DNA has been shown to enter the cytoplasm where it binds to "DNA sensors," which in turn initiate signaling cascades. Here we report data that support a novel signaling pathway in response to oxidative stress mediated by specific guanine-rich sequences that can fold into G-quadruplex DNA (G4DNA). In response to oxidative stress, we demonstrate that sequences capable of forming G4DNA appear at increasing levels in the cytoplasm and participate in assembly of stress granules. Identified proteins that bind to endogenous G4DNA in the cytoplasm are known to modulate mRNA translation and participate in stress granule formation. Consistent with these findings, stress granule formation is known to regulate mRNA translation during oxidative stress. We propose a signaling pathway whereby cells can rapidly respond to DNA damage caused by oxidative stress. Guanine-rich sequences that are excised from damaged genomic DNA are proposed to enter the cytoplasm where they can regulate translation through stress granule formation. This newly proposed role for G4DNA provides an additional molecular explanation for why such sequences are prevalent in the human genome.

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Section: Resultsmentioning

confidence: 99%

Evidence That G-quadruplex DNA Accumulates in the Cytoplasm and Participates in Stress Granule Assembly in Response to Oxidative Stress

Byrd¹,

Zybailov

Maddukuri³

et al. 2016

Journal of Biological Chemistry

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“…Although GQ RNA shape-based recognition is not well established, it is known that other protein domains can also preferentially recognize GQ-forming RNAs including: the arginine-glycine-glycine repeat (RGG) domain of Fragile-X mental retardation protein (FMRP), the glycine-argininerich (GAR) domain of TRF2, and the N-terminal domain of the DEAH-box ATP-dependent helicase 36 (DHX36) (Deng et al 2009;Meier et al 2013;Chen et al 2015;Vasilyev et al 2015). With the development of GQ-specific antibodies that have unambiguously identified GQ TERRA RNA structures in living cells (Xu et al 2010;Di Antonio et al 2012;Biffi et al 2013Biffi et al , 2014, more biophysical studies are needed that probe how protein domains and multidomain protein complexes recognize the molecular architecture of GQ RNAs.…”

Section: Discussionmentioning

confidence: 99%

G-quadruplex RNA binding and recognition by the lysine-specific histone demethylase-1 enzyme

Hirschi

Martin

Luka

et al. 2016

RNA

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Lysine-specific histone demethylase 1 (LSD1) is an essential epigenetic regulator in metazoans and requires the co-repressor element-1 silencing transcription factor (CoREST) to efficiently catalyze the removal of mono-and dimethyl functional groups from histone 3 at lysine positions 4 and 9 (H3K4/9). LSD1 interacts with over 60 regulatory proteins and also associates with lncRNAs (TERRA, HOTAIR), suggesting a regulatory role for RNA in LSD1 function. We report that a stacked, intramolecular G-quadruplex (GQ) forming TERRA RNA (GG[UUAGGG] 8 UUA) binds tightly to the functional LSD1-CoREST complex (K d ≈ 96 nM), in contrast to a single GQ RNA unit ([UUAGGG] 4 U), a GQ DNA ([TTAGGG] 4 T), or an unstructured single-stranded RNA. Stabilization of a parallel-stranded GQ RNA structure by monovalent potassium ions (K + ) is required for high affinity binding to the LSD1-CoREST complex. These data indicate that LSD1 can distinguish between RNA and DNA as well as structured versus unstructured nucleotide motifs. Further, cross-linking mass spectrometry identified the primary location of GQ RNA binding within the SWIRM/amine oxidase domain (AOD) of LSD1. An ssRNA binding region adjacent to this GQ binding site was also identified via X-ray crystallography. This RNA binding interface is consistent with kinetic assays, demonstrating that a GQ-forming RNA can serve as a noncompetitive inhibitor of LSD1-catalyzed demethylation. The identification of a GQ RNA binding site coupled with kinetic data suggests that structured RNAs can function as regulatory molecules in LSD1-mediated mechanisms.

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Section: Mechanism Of G4 Disruption By Dhx36mentioning

confidence: 99%

“…The disruption rate was shown to be greater for a G4 with fewer G-quartets, suggesting that less stable G4s are disrupted more efficiently by DHX36 (26). A small Nterminal domain directs DHX36 to G4s by binding specifically to these structures (28)(29)(30).Nevertheless, important aspects of the mechanism of G4 disruption by DHX36 remain unknown. DEAH/RHA family helicases (abbreviated herein as DEAH family) typically use a translocation-based mechanism for unwinding DNA or RNA helices, a mechanism in which ATP drives directional motion along a sequestered single strand, displacing the complementary strand (31).…”

mentioning

confidence: 99%

The G-quadruplex (G4) resolvase DHX36 efficiently and specifically disrupts DNA G4s via a translocation-based helicase mechanism

Yangyuoru

Bradburn

et al. 2018

Journal of Biological Chemistry

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Single-stranded DNA (ssDNA) and RNA regions that include at least four closely spaced runs of three or more consecutive guanosines strongly tend to fold into stable G-quadruplexes (G4s). G4s play key roles as DNA regulatory sites and as kinetic traps that can inhibit biological processes, but how G4s are regulated in cells remains largely unknown. Here, we developed a kinetic framework for G4 disruption by DEAH-box helicase 36 (DHX36), the dominant G4 resolvase in human cells. Using tetramolecular DNA and RNA G4s with four to six G-quartets, we found that DHX36-mediated disruption is highly efficient, with rates that depend on G4 length under saturating conditions (k cat ) but not under subsaturating conditions (k cat /K M ). These results suggest that a step during G4 disruption limits the k cat value and that DHX36 binding limits k cat /K M . Similar results were obtained for unimolecular DNA G4s. DHX36 activity depended on a 3′ ssDNA extension and was blocked by a polyethylene glycol linker, indicating that DHX36 loads onto the extension and translocates 3′-5′ toward the G4. DHX36 unwound dsDNA poorly compared with G4s of comparable intrinsic lifetime. Interestingly, we observed that DHX36 has striking 3′-extension sequence preferences that differ for G4 disruption and dsDNA unwinding, most likely arising from differences in the rate-limiting step for the two activities. Our results indicate that DHX36 disrupts G4s with a conventional helicase mechanism that is tuned for great efficiency and specificity for G4s. The dependence of DHX36 on the 3′-extension sequence suggests that the extent of formation of genomic G4s may not track directly with G4 stability. ___________________________________Single-stranded DNA and RNA segments that include at least four closely-spaced runs of three or more consecutive guanosine nucleotides have a strong intrinsic propensity to fold into stable G-quadruplex structures (G4s) (1) (Fig. 1A, top left). The genomes of humans and many other eukaryotes are replete with putative G4-forming sequences, and pronounced, conserved patterns have been observed in their distribution, suggesting that Gquadruplex structures form at least transiently in cells and perform conserved functions (2-6). Supporting this hypothesis, G4s have been detected in cells for both .To function as regulatory elements, G4s must be folded and disrupted in a regulated way. In addition, processes that require single-stranded DNA or RNA, such as replication and translation, require that G4s be temporarily disrupted. The high stability and long intrinsic lifetime of G4s suggest Mechanism of G4 disruption by DHX362 that their efficient disruption requires the activity of ATP-dependent enzymes such as helicases.Supporting this view, incorporation of sequences predicted to form particularly stable G4s leads to genetic instability in yeast, suggesting that these G4 structures are not processed efficiently and pose blocks to replication (12). Indeed, several helicases have been shown to possess G4 disruption acti...

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Binding of G-quadruplexes to the N-terminal Recognition Domain of the RNA Helicase Associated with AU-rich Element (RHAU)

Cited by 53 publications

References 49 publications

Evidence That G-quadruplex DNA Accumulates in the Cytoplasm and Participates in Stress Granule Assembly in Response to Oxidative Stress

Evidence That G-quadruplex DNA Accumulates in the Cytoplasm and Participates in Stress Granule Assembly in Response to Oxidative Stress

G-quadruplex RNA binding and recognition by the lysine-specific histone demethylase-1 enzyme

The G-quadruplex (G4) resolvase DHX36 efficiently and specifically disrupts DNA G4s via a translocation-based helicase mechanism

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