Negative Regulation of AAA+ ATPase Assembly by Two Component Receiver Domains: A Transcription Activation Mechanism that is Conserved in Mesophilic and Extremely Hyperthermophilic Bacteria

Doucleff, Michaeleen; Chen, Baoyu; Maris, A.E.; Wemmer, David E.; Kondrashkina, Elena; Nixon, B. Tracy

doi:10.1016/j.jmb.2005.08.003

Cited by 56 publications

(97 citation statements)

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“…The structural model that we describe for activated NtrC, in which the activated receiver domain of one subunit is in contact with the ATPase domain of a second one to stabilize ring assembly, differs dramatically from the model proposed to explain how two-component signal transduction regulates assembly of AAA+ ATPase domains in the enhancer-binding proteins NtrC1 and DctD Doucleff et al 2005). Consequently, the new model establishes the structural differences underlying positive versus negative regulation for this family of enhancer-binding proteins.…”

Section: Discussionmentioning

confidence: 71%

“…By the time P i has been released (bottom), the GAFTGA region has let go of 54 and become disordered (bottom right) and the DNA-binding regions detached from the central ring (bottom left), releasing their constraint on the DNA. mechanism, deletions that truncate or remove the receiver domain result in constitutively active proteins (see Doucleff et al 2005). In stark contrast, positively regulated activators like NtrC require the presence of the phosphorylated receiver domain to function in ATPase and transcription activation assays (Drummond et al 1990;Weiss et al 1992a,b;Klose et al 1993).…”

Section: Mechanism Of Positive Activation In 54 -Dependent Aaa+ Atpasmentioning

confidence: 99%

“…In their inactive states these proteins are usually dimers that bind to pairs of tandem sites in the enhancer elements. Upon activation via the regulatory domains, they oligomerize into ATPase-active rings that use the energy from ATP hydrolysis to physically remodel closed complexes of 54 holoenzyme and promoter DNA (Rombel et al 1998;Neuwald et al 1999;Chaney et al 2001;Lee et al 2003).Crystal or NMR structures are available for several isolated domains and truncation constructs of EBPs including NtrC, DctD, PspF, ZraR, and NtrC1 (Volkman et al 1995;Kern et al 1999;Pelton et al 1999;Meyer et al 2001;Park et al 2002;Hastings et al 2003;Lee et al 2003;Doucleff et al 2005;Rappas et al 2005;Sallai and Tucker 2005). Existing structural information on DctD and NtrC1 has been used to provide a model of how two-component signal transduction can regulate assembly of their AAA+ ATPase domains ), but this model fails to explain regulation in the closely related protein NtrC (40% sequence identity, 60% sequence similarity) Hastings et al 2003).…”

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confidence: 99%

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The structural basis for regulated assembly and function of the transcriptional activator NtrC

Carlo¹,

Chen²,

Hoover³

et al. 2006

Genes Dev.

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In two-component signal transduction, an input triggers phosphorylation of receiver domains that regulate the status of output modules. One such module is the AAA+ ATPase domain in bacterial enhancer-binding proteins that remodel the 54 form of RNA polymerase. We report X-ray solution scattering and electron microscopy structures of the activated, full-length nitrogen-regulatory protein C (NtrC) showing a novel mechanism for regulation of AAA+ ATPase assembly via the juxtaposition of the receiver domains and ATPase ring. Accompanying the hydrolysis cycle that is required for transcriptional activation, we observed major order-disorder changes in the GAFTGA loops involved in 54 binding, as well as in the DNA-binding domains. The 54 holoenzyme form of RNA polymerase forms closed DNA complexes that open only with the help of enhancer-binding proteins (EBPs). For the nitrogen-regulatory protein C of enteric bacteria (NtrC), such activity starts a cascade of events that may ultimately lead to the activation of transcription for as much as 2% of the genome (Zimmer et al. 2000). Sequence analysis of >160054 -dependent transcriptional activators shows that in addition to their oligomerization/AAA+ ATPase domain, they also have a DNA-binding domain and a regulatory domain, which in 50% of cases is a twocomponent receiver domain (Bateman et al. 2004). The helix-turn-helix DNA-binding domain recognizes enhancer-like sequences between 100 and 150 bp upstream of the promoter. In their inactive states these proteins are usually dimers that bind to pairs of tandem sites in the enhancer elements. Upon activation via the regulatory domains, they oligomerize into ATPase-active rings that use the energy from ATP hydrolysis to physically remodel closed complexes of 54 holoenzyme and promoter DNA (Rombel et al. 1998;Neuwald et al. 1999;Chaney et al. 2001;Lee et al. 2003).Crystal or NMR structures are available for several isolated domains and truncation constructs of EBPs including NtrC, DctD, PspF, ZraR, and NtrC1 (Volkman et al. 1995;Kern et al. 1999;Pelton et al. 1999;Meyer et al. 2001;Park et al. 2002;Hastings et al. 2003;Lee et al. 2003;Doucleff et al. 2005;Rappas et al. 2005;Sallai and Tucker 2005). Existing structural information on DctD and NtrC1 has been used to provide a model of how two-component signal transduction can regulate assembly of their AAA+ ATPase domains ), but this model fails to explain regulation in the closely related protein NtrC (40% sequence identity, 60% sequence similarity) Hastings et al. 2003). Here we report both SAXS/WAXS (small-and wide-angle X-ray scattering) and EM (electron microscopy) structures of the full-length, activated form of NtrC from Salmonella typhimurium. Docking of atomic models for the individual domains into the structures reveals their organization within the activated ring and uncovers a novel mechanism for the use of two-compo-

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Section: Discussionmentioning

confidence: 71%

Section: Mechanism Of Positive Activation In 54 -Dependent Aaa+ Atpasmentioning

confidence: 99%

mentioning

confidence: 99%

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The structural basis for regulated assembly and function of the transcriptional activator NtrC

Carlo¹,

Chen²,

Hoover³

et al. 2006

Genes Dev.

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“…The N-terminal domain controls the ability of the EBP to form hexameric or heptameric oligomers in response to a particular stimulus (Lee et al, 1994;Chen et al, 2008;Lee et al, 2003;Doucleff et al, 2005a;De Carlo et al, 2006). The EBPs can be classified by the type of domain that is present in the N-terminal region (Studholme & Dixon, 2003).…”

Section: Introductionmentioning

confidence: 99%

Identification of the binding site of the σ 54 hetero-oligomeric FleQ/FleT activator in the flagellar promoters of Rhodobacter sphaeroides

et al. 2009

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Expression of the flagellar genes in Rhodobacter sphaeroides is dependent on one of the four sigma-54 factors present in this bacterium and on the enhancer binding proteins (EBPs) FleQ and FleT. These proteins, in contrast to other well-characterized EBPs, carry out activation as a hetero-oligomeric complex. To further characterize the molecular properties of this complex we mapped the binding sites or upstream activation sequences (UASs) of six different flagellar promoters. In most cases the UASs were identified at approximately 100 bp upstream from the promoter. However, the activity of the divergent promoters flhAp-flgAp, which are separated by only 53 bp, is mainly dependent on a UAS located approximately 200 bp downstream from each promoter. Interestingly, a significant amount of activation mediated by the upstream or contralateral UAS was also detected, suggesting that the architecture of this region is important for the correct regulation of these promoters. Sequence analysis of the regions carrying the potential FleQ/FleT binding sites revealed a conserved motif. In vivo footprinting experiments with the motAp promoter allowed us to identify a protected region that overlaps with this motif. These results allow us to propose a consensus sequence that represents the binding site of the FleQ/ FleT activating complex. INTRODUCTIONPromoter recognition in bacteria is mediated by the RNA polymerase core (E) associated with a sigma factor. There are two families of sigma factors, the s 70 family and that of s 54 . These two families differ not only in their sequence but also in the mechanism that brings about transcriptional initiation. Whereas Es 70 is capable of forming open complex by itself, Es 54 requires an activator protein that, through ATP hydrolysis, allows open complex formation (for reviews, see Buck et al., 2000;Wigneshweraraj et al., 2005). Activator proteins of Es 54 usually bind approximately 100 bp upstream of the promoter sequence, and a DNA loop favours the interaction of the activator with the RNA polymerase holoenzyme bound to the promoter (Reitzer & Magasanik, 1986;Ninfa et al., 1987;Buck et al., 1986;Hoover et al., 1990;Su et al., 1990;Wedel et al., 1990). The mechanism that allows open complex formation is still unknown, but the current model suggests that the activator protein remodels the nucleoprotein complex formed by Es 54 and DNA in order to initiate transcription Cannon et al., 2000;Burrows et al., 2004;Rappas et al., 2007). The DNA region to which the activator proteins bind is known as the upstream activation sequence or UAS (Buck et al., 1986;Morett et al., 1988). It has been shown for the glnA promoter that this region can be moved more than 1000 bp without losing its ability to stimulate transcription (Ninfa et al., 1987). Based on this property, the UASs are also known as enhancers, and the activator proteins as enhancer binding proteins, or EBPs.The s 54 -dependent promoters show a highly conserved consensus sequence, and their main characteristics are the dinucleotides GG ...

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“…Indeed, it was in an attempt to form such a complex with beryllofluoride and ADP at the ATP binding site of the 54 activator NtrC that Dalai Yan et al found activation of the protein rather than the anticipated inhibition (15). Subsequent detailed investigation showed that addition of BeF 3 Ϫ yielded an activity level as high as that from phosphorylation in a direct assay of transcriptional activation by NtrC. The interaction with BeF 3 Ϫ is very specific to the active site in the response regulator receiver domain, which is normally activated by phosphotransfer from a histidine kinase as part of a two-component signaling system (14).…”

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confidence: 99%

Negative Regulation of AAA+ ATPase Assembly by Two Component Receiver Domains: A Transcription Activation Mechanism that is Conserved in Mesophilic and Extremely Hyperthermophilic Bacteria

Cited by 56 publications

References 46 publications

The structural basis for regulated assembly and function of the transcriptional activator NtrC

The structural basis for regulated assembly and function of the transcriptional activator NtrC

Identification of the binding site of the σ 54 hetero-oligomeric FleQ/FleT activator in the flagellar promoters of Rhodobacter sphaeroides

Beryllofluoride Binding Mimics Phosphorylation of Aspartate in Response Regulators

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