Still Looking for the Magic Spot: The Crystallographically Defined Binding Site for ppGpp on RNA Polymerase Is Unlikely to Be Responsible for rRNA Transcription Regulation

Vrentas, Catherine E.; Gaál, Tamás; Berkmen, Melanie B.; Rutherford, Steven T.; Haugen, Shanil P.; Ross, Wilma; Gourse, Richard L.

doi:10.1016/j.jmb.2008.01.042

Cited by 75 publications

(82 citation statements)

References 45 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…RelA/SpoT homologs are found throughout the bacterial kingdom (5), although the mechanism of ppGpp action on rRNA transcription appears to be indirect in some cases (10,42). DksA has been studied far less than ppGpp in organisms other than E. coli.…”

Section: Discussionmentioning

confidence: 99%

“…DksA has been studied far less than ppGpp in organisms other than E. coli. Sequence homologs of DksA are present throughout proteobacteria, but they are not obvious in many other species, including some Gram-negative or some Gram-positive thermophiles (10,42). However, structurally similar proteins could be present but not recognized in sequence comparisons.…”

Section: Discussionmentioning

confidence: 99%

“…ppGpp binds directly to E. coli RNA polymerase (RNAP) and inhibits transcription from rRNA promoters (9), although the identity of the ppGpp binding site on RNAP remains unclear (10). However, for ppGpp to exert its full effect on transcription, RNAP has to be modified by the small protein, DksA (11,12).…”

mentioning

confidence: 99%

See 2 more Smart Citations

Direct regulation of Escherichia coli ribosomal protein promoters by the transcription factors ppGpp and DksA

Lemke

Sanchez-Vazquez²,

Burgos³

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

140

144

View full text Add to dashboard Cite

We show here that the promoters for many of the Escherichia coli ribosomal protein operons are regulated directly by two transcription factors, the small RNA polymerase-binding protein DksA and the nutritional stress-induced nucleotide ppGpp. ppGpp and DksA work together to inhibit transcription initiation from ribosomal protein promoters in vitro and in vivo. The degree of promoter regulation by ppGpp/DksA varies among the r-protein promoters, but some are inhibited almost as much as rRNA promoters. Thus, many r-protein operons are regulated at the level of transcription in addition to their control by the classic translational feedback systems discovered ∼30 y ago. We conclude that direct control of r-protein promoters and rRNA promoters by the same signal, ppGpp/DksA, makes a major contribution to the balanced and coordinated synthesis rates of all of the ribosomal components.ribosome synthesis | transcriptional control | stringent response | translational control P rotein synthesis is the major consumer of cellular energy in bacteria. Because the number of ribosomes is the primary determinant of the level of translation, and ribosome synthesis itself is an energy-intensive process, there are mechanisms that prevent over-or underinvestment of cellular resources in ribosome synthesis (1). Both ribosomal RNA (rRNA) and ribosomal protein (r-protein) synthesis rates are thereby tightly regulated in Escherichia coli (for reviews, see refs. 2, 3).One of the earliest reported examples of regulation of bacterial ribosome synthesis is a stress response referred to as "the stringent response," in which rRNA transcription is inhibited in cells starved for amino acids or some other nutrients (4). In this response, uncharged tRNAs induce the ribosome-associated RelA and/or SpoT proteins to synthesize ppGpp (5-7). [The term "ppGpp" is used here to describe both the unusual nucleotide guanosine-3′,5′-(bis)pyrophosphate and its pentaphosphate precursor.] ppGpp concentrations change not only after complete starvations but also after less severe shifts in nutritional conditions, coordinating rRNA synthesis with the need for protein synthesis. Shifts to a more favorable nutritional condition result in a decrease in the concentration of ppGpp and a corresponding increase in rRNA promoter activity, whereas shifts to a less favorable condition result in an increase in ppGpp and a corresponding decrease in rRNA transcription (8).ppGpp binds directly to E. coli RNA polymerase (RNAP) and inhibits transcription from rRNA promoters (9), although the identity of the ppGpp binding site on RNAP remains unclear (10). However, for ppGpp to exert its full effect on transcription, RNAP has to be modified by the small protein, DksA (11,12). Unlike ppGpp, DksA is present at high concentrations in cells under all conditions that have been examined (11,13). rRNA transcription initiation is also regulated by the concentration of the first nucleotide in the transcript (8,14) and by at least one DNA binding factor, the 11.2-kDa Fis protein (15). Tog...

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Direct regulation of Escherichia coli ribosomal protein promoters by the transcription factors ppGpp and DksA

Lemke

Sanchez-Vazquez²,

Burgos³

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

140

144

View full text Add to dashboard Cite

show abstract

“…ppGpp) and antibiotics (e.g. lipiarmycin) that specifically affect E. coli but not the Thermus RNAPs (20,21). These structural insights are important to identify their binding sites and to understand the mechanisms of action.…”

Section: Rna Polymerase (Rnap)mentioning

confidence: 99%

X-ray Crystal Structure of Escherichia coli RNA Polymerase σ70 Holoenzyme

Murakami

2013

Journal of Biological Chemistry

184

221

View full text Add to dashboard Cite

Background: A crystal structure of Escherichia coli RNA polymerase (RNAP) has not been determined. Results: The 1.1 and ␣ subunit C-terminal domain structures have been determined in the context of an intact RNAP. Conclusion: 1.1 localizes within the RNAP DNA-binding channel and must disengage from this site to form an open complex. Significance: This work enables future structure determination of bacterial RNAP mutants.

show abstract

“…Additional detailed studies suggest that DksA, which binds to the secondary channel of RNA polymerase, interferes allosterically with the transcription initiation site, affecting the transition from Combined effects of ppGpp, DksA and the iNTPs during stringent control Both ppGpp and DksA exert their effect by direct binding to the secondary channel of RNA polymerase (Artsimovitch et al, 2004;Perederina et al, 2004). Both molecules are directly located near the active centre, where nucleotides are incorporated, although some of the conclusions with respect to ppGpp and E. coli RNA polymerase have been challenged by a controversial study (Vrentas et al, 2008). It is known that binary open (RP o ) complexes of the stringent-sensitive rRNA P1 promoters are intrinsically unstable, and only after binding of the first substrate NTP are stable kinetic intermediates (RP i ) formed (Barker et al, 2001;Gourse, 1988;Langert et al, 1991).…”

Section: Discussionmentioning

confidence: 99%

Differential stringent control of Escherichia coli rRNA promoters: effects of ppGpp, DksA and the initiating nucleotides

et al. 2011

View full text Add to dashboard Cite

Transcription of rRNAs in Escherichia coli is directed from seven redundant rRNA operons, which are mainly regulated by their P1 promoters. Here we demonstrate by in vivo measurements that the amounts of individual rRNAs transcribed from the different operons under normal growth vary noticeably although the structures of all the P1 promoters are very similar. Moreover, we show that starvation for amino acids does not affect the seven P1 promoters in the same way. Notably, reduction of transcription from rrnD P1 was significantly lower compared to the other P1 promoters. The presence of DksA was shown to be crucial for the ppGpp-dependent downregulation of all P1 promoters. Because rrnD P1 is the only rrn promoter starting with GTP instead of ATP, we performed studies with a mutant rrnD promoter, where the initiating G+1 is replaced by A+1. These analyses demonstrated that the ppGpp sensitivity of rrn P1 promoters depends on the nature and concentration of initiating nucleoside triphosphates (iNTPs). Our results support the notion that the seven rRNA operons are differentially regulated and underline the importance of a concerted activity between ppGpp, DksA and an adequate concentration of the respective iNTP. INTRODUCTIONThe rapid adaptation of cell growth in response to changing environmental conditions is central for the fitness and survival of bacteria. Bacterial growth rate is largely determined by the number of ribosomes. Hence, regulation of the translational components plays a central role in adaptation. The biogenesis of ribosomes, representing one of the cell's most energy-intensive activities, is governed by a complex network of regulation (Schneider et al., 2003;Wagner, 2009). The key molecules in this network are rRNAs. In contrast, the synthesis of ribosomal proteins, despite having similar regulatory mechanisms to those for rRNA, is largely linked to rRNA transcription by a translational feedback mechanism, which couples ribosomal protein translation to the amount of available rRNAs (Keener & Nomura, 1996;Lemke et al., 2009;Nomura et al., 1984). Therefore, the precise adjustment of rRNA synthesis rate to the nutritional quality and supply from the environment ensures that an appropriate number of ribosomes will be produced and energy resources are not wasted (Condon et al., 1995b;Wagner, 1994Wagner, , 2009).In Escherichia coli, rRNA genes (16S, 23S and 5S rRNA), together with at least one tRNA gene, are organized in a redundant fashion in seven different transcriptional units (the rrnA, rrnB, rrnC, rrnD, rrnE, rrnG and rrnH operons). Although the seven ribosomal operons exhibit a high structural similarity, there are several striking sequence micro-heterogeneities, which could potentially contribute to functionally different ribosome populations (Wagner, 2009). Evidence for such specialized ribosomes in various organisms has been presented (Gunderson et al., 1987;Kim et al., 2007; Ló pez-Ló pez et al., 2007). If the different rRNA operons encode functionally distinct molecules one would expect tha...

show abstract

Still Looking for the Magic Spot: The Crystallographically Defined Binding Site for ppGpp on RNA Polymerase Is Unlikely to Be Responsible for rRNA Transcription Regulation

Cited by 75 publications

References 45 publications

Direct regulation of Escherichia coli ribosomal protein promoters by the transcription factors ppGpp and DksA

Direct regulation of Escherichia coli ribosomal protein promoters by the transcription factors ppGpp and DksA

X-ray Crystal Structure of Escherichia coli RNA Polymerase σ70 Holoenzyme

Differential stringent control of Escherichia coli rRNA promoters: effects of ppGpp, DksA and the initiating nucleotides

Contact Info

Product

Resources

About