Essential Steps in the ppGpp-dependent Regulation of Bacterial Ribosomal RNA Promoters Can Be Explained by Substrate Competition

Abstract

Transcription of stable RNA genes is known to be dramatically reduced in the presence of guanosine tetraphosphate (ppGpp), the mediator of the stringent response. Using in vitro transcription systems with ribosomal RNA P1 promoters, we have analyzed which step of the initiation cycle is inhibited by the effector ppGpp. We show that formation of the ternary transcription initiation complex consisting of RNA polymerase holoenzyme, the promoter DNA, and the first initiating nucleotide triphosphate is the major step at which ppGpp exerts its regulation. Neither primary binding of RNA polymerase to the promoter nor isomerization to the open binary complexes or the subsequent promoter clearance steps contributes notably to the observed inhibition. The effect of ppGpp-dependent inhibition in the formation of the ternary transcription initiation complex could be mimicked by nucleotide derivatives known to bind to the RNA polymerase active center. Using these model compounds, almost identical inhibition characteristics were observed as seen with ppGpp. The results support the previously published model, which suggests that ppGpp-dependent inhibition is based on competition between the inhibitor molecules and NTP substrates for access to the active center of RNA polymerase. Transcription of stable RNA genes is known to be dramatically reduced in the presence of guanosine tetraphosphate (ppGpp), the mediator of the stringent response. Using in vitro transcription systems with ribosomal RNA P1 promoters, we have analyzed which step of the initiation cycle is inhibited by the effector ppGpp. We show that formation of the ternary transcription initiation complex consisting of RNA polymerase holoenzyme, the promoter DNA, and the first initiating nucleotide triphosphate is the major step at which ppGpp exerts its regulation. Neither primary binding of RNA polymerase to the promoter nor isomerization to the open binary complexes or the subsequent promoter clearance steps contributes notably to the observed inhibition. The effect of ppGpp-dependent inhibition in the formation of the ternary transcription initiation complex could be mimicked by nucleotide derivatives known to bind to the RNA polymerase active center. Using these model compounds, almost identical inhibition characteristics were observed as seen with ppGpp. The results support the previously published model, which suggests that ppGpp-dependent inhibition is based on competition between the inhibitor molecules and NTP substrates for access to the active center of RNA polymerase. ribosomal RNA RNA polymerase promoter closed complex binary open complex ternary open complex or initiating complex that contains at least one bound substrate NTP elongating complex Bacterial cells adapt their growth very efficiently in response to changes in the environmental conditions. The cellular growth is generally determined by the capacity for protein synthesis, which itself is tightly linked to the number of ribosomes. During amino acid deprivation bacterial cells react, among other metabolic adaptations, by an immediate transcriptional shut-down of ribosomal RNAs (rRNAs),1 the key molecules for the constitution of the protein synthesis apparatus. This global regulation is triggered by the rapid accumulation of the effector molecule guanosine tetraphosphate (ppGpp) and termed the stringent response (1Cashel M. Gentry D.R. Hernandez V.J. Vinella D. Neidhard F.C. Curtiss III, R. Ingraham J.L. Lin E.C.C. Low K.B. Magasanik B. Reznikoff W.S. Riley M. Schaechter M. Umbarger H.E. The Stringent Response: Escherichia coli and Salmonella Cellular and Molecular Biology. 2. ASM Press, Washington, D. C.1996: 1458-1496Google Scholar). Although gradual progress in understanding this remarkable network has been made in recent years, the molecular details of the regulatory mechanism(s) are still obscure (2Barker M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: 689-702Crossref PubMed M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed M. R. J. PubMed M. R. J. M. R. J. Biol. PubMed Hernandez V.J. J. Biol. 2001; PubMed D. Mol. PubMed M. T. PubMed M. J. PubMed Scholar). The that amino acid are the stringent response and that in and in vitro have results have understanding of the of ppGpp-dependent regulation R. Transcription in Press, Scholar). ppGpp-dependent regulation is to the steps of transcription initiation has been to the of RNA polymerase are M. R. J. Biol. PubMed J. PubMed is that RNA polymerase is the for the effector molecule ppGpp Hernandez V.J. J. Biol. 2001; PubMed D. Mol. PubMed PubMed Scholar). has been to to the active center by the and The presence of ppGpp the of the in that transcription stringent and is in the RNA polymerase have been which polymerase and initiation of RNA polymerase at ppGpp V.J. M. J. Mol. Biol. PubMed Scholar). This has been observed for other and as M. T. PubMed T. J. Biol. PubMed is or is by its promoter is known for that stringent regulation of the promoter J. PubMed Scholar). This is termed the and has been to be for stringent regulation. is that other of in to the are in transcription is stringent or M. R. J. Scholar). in the have that ppGpp-dependent are by of their binary open the stable open complexes in presence of their M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed M. R. J. PubMed R.L. PubMed T. Gourse R.L. PubMed Scholar). the of the of open binary complexes is very and complexes have to at NTP formation of the ternary complexes be the step for transcription and at which step of the initiation ppGpp transcription this we have analyzed the effect of ppGpp on the steps to transcription initiation at P1 were and the promoter as We show that ppGpp the at the the binary open complexes to the ternary initiation The of the substrate in this that competition between the substrate and ppGpp for the is This is by almost identical inhibition characteristics ppGpp is by NTP substrate known to bind to the active of RNA this we have to of the that are still with to the of the stringent This of regulation is by the effector molecule which transcription of in stable RNA The of this global changes in response to of environmental for which amino acid is the to the of this regulatory of bacterial we still are are and in is for that RNA polymerase is the for ppGpp Hernandez V.J. J. Biol. 2001; PubMed D. Mol. PubMed PubMed the are is known that the promoter is stringent or M. R. J. PubMed J. J. PubMed M. R. PubMed Scholar). and of transcription are by be effect of the ribosomal RNA the for stringent for the protein or amino acid are known to be at ppGpp of the recent that the of genes is by (2Barker M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: 689-702Crossref PubMed have been H.E. J. Biol. PubMed PubMed J. PubMed the of the stringent steps of the transcription cycle have been to be for regulation. formation of the open promoter complex has been as the step for the J. Biol. PubMed D. PubMed Scholar). in the closed complex by inhibition at promoter by of and promoter M. R. J. PubMed Scholar). The model is with the model, which suggests that RNA polymerase in with of the is triggered by bound ppGpp. RNA polymerase ppGpp stable RNA promoters, the with bound ppGpp J. Biol. PubMed Scholar). on the that stable RNA open for and stringent have been M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed M.M. Gourse R.L. J. 2001; PubMed Scholar). to ppGpp has effect on stringent stable RNA stringent regulation is as effect of RNA polymerase which results transcriptional shut-down of the stable RNA in and in vitro for transcriptional inhibition by ppGpp at the promoter Hernandez V.J. J. Biol. PubMed J. PubMed Scholar). the promoter stable open complexes that ppGpp-dependent regulation is to with open based on for stringent have been previously PubMed Ingraham J. J. J. Mol. Biol. PubMed Scholar). The and the that by itself the that is one for the stringent This is by the that ppGpp is to steps transcription initiation M. R. J. Biol. PubMed J. PubMed J. Biol. PubMed this we have in to which of the initiation are by ppGpp and to the molecular by which ppGpp exerts regulatory We the ribosomal RNA P1 the and which in their nucleotide for and for in the model, which that the NTP in the the of stringent T. Gourse R.L. PubMed Scholar). we the which stringent and which is known to be by ppGpp. with the we very inhibition in transcription the stable RNA were inhibited the conditions. the ppGpp inhibition of the and P1 promoter inhibition with for this is by the of the promoter P1 in and in the or the the that the major effect of ppGpp is or the formation of the ternary transcription initiation This be the inhibition is observed ppGpp is or at the formation of the ternary complex the initiation cycle initiation or to inhibition we show that the of initiating is for the of inhibition. NTP ppGpp-dependent inhibition We could that the the open complex is very in presence of ppGpp. We this as that the of ternary transcription complexes are dramatically by ppGpp with M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed we that the in of the ternary complexes in presence or of ppGpp is to the of inhibition we that the of ternary complex formation is reduced in presence of ppGpp. This effect be by an in the of the initiating these results are with previously published model, that ppGpp with the initiating NTP substrates R. J. Mol. Scholar). The model the that the for substrate to RNA polymerase active center is the by of the and M. PubMed Scholar). The of this nucleotide is to of one NTP at to ppGpp. is that at ppGpp of substrate to the active is This the for substrate binding are and the complexes are the for substrate are as is the for initiation complexes at stable RNA promoters, the competition Stringent be by the promoter which its and with RNA the of the initiating complex and the for the first substrate The effector molecule which to the stringent competition to the ppGpp with the substrate for access to the active center of RNA polymerase. or initiation complexes are are in an to previously published J. Biol. PubMed that ppGpp the of RNA polymerase by of the transcription The by that ppGpp-dependent inhibition to be with the triphosphate are for this the of ppGpp-dependent inhibition on be that on ternary initiation complex formation the promoter is The were at with transcription complexes at the and were is the to the complex with on The observed inhibition be the of one inhibition and inhibition be complex substrate competition is in the of ppGpp-dependent inhibition by the effector molecule ppGpp with nucleotide We and that are known to be as and are to the active center of RNA polymerase. be seen in and these inhibited transcription of the stable RNA almost the characteristics of inhibition and were at the as ppGpp. in the promoter for the or at the step to formation of the ternary initiation the on the of the as observed with the reduced the at which the ternary complexes are We that these are an we these as that substrate competition major in the of stringent is the that substrates other of ppGpp-dependent inhibition. substrates with ppGpp binding and in the as be the for ppGpp. recent in initiation the at the that inhibition Hernandez V.J. J. Biol. PubMed Scholar). the of this the that ppGpp formation of the first and this step is by the that the effect of ppGpp is in the presence of the The as to this the model and to the of inhibition or with are is for the regulation of NTP access to the active of RNA and the for of NTP substrates has been in the that is for the have been the RNA the by and or are to for RNA polymerase The with the at the active of RNA polymerase. The the and the for the active This termed by the of the and the of substrate Mol. PubMed of competition on RNA polymerase the complex be changes of the transcription are in of the that RNA polymerase be by as in the of the the of the is by in the molecule binding to stringent or the with an effector molecule ppGpp the of the could of the stringent which to be substrate have that ppGpp the of with RNA polymerase M. T. PubMed T. J. Biol. PubMed T. J. Biol. PubMed Scholar). this ppGpp could be an effector that at the and which the complex of the that is known to with the PubMed PubMed Scholar). Bacterial cells adapt their growth very efficiently in response to changes in the environmental conditions. The cellular growth is generally determined by the capacity for protein synthesis, which itself is tightly linked to the number of ribosomes. During amino acid deprivation bacterial cells react, among other metabolic adaptations, by an immediate transcriptional shut-down of ribosomal RNAs (rRNAs),1 the key molecules for the constitution of the protein synthesis apparatus. This global regulation is triggered by the rapid accumulation of the effector molecule guanosine tetraphosphate (ppGpp) and termed the stringent response (1Cashel M. Gentry D.R. Hernandez V.J. Vinella D. Neidhard F.C. Curtiss III, R. Ingraham J.L. Lin E.C.C. Low K.B. Magasanik B. Reznikoff W.S. Riley M. Schaechter M. Umbarger H.E. The Stringent Response: Escherichia coli and Salmonella Cellular and Molecular Biology. 2. ASM Press, Washington, D. C.1996: 1458-1496Google Scholar). Although gradual progress in understanding this remarkable network has been made in recent years, the molecular details of the regulatory mechanism(s) are still obscure (2Barker M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: 689-702Crossref PubMed M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed M. R. J. PubMed M. R. J. M. R. J. Biol. PubMed Hernandez V.J. J. Biol. 2001; PubMed D. Mol. PubMed M. T. PubMed M. J. PubMed Scholar). The that amino acid are the stringent response and that in and in vitro have results have understanding of the of ppGpp-dependent regulation R. Transcription in Press, Scholar). ppGpp-dependent regulation is to the steps of transcription initiation has been to the of RNA polymerase are M. R. J. Biol. PubMed J. PubMed Scholar). is that RNA polymerase is the for the effector molecule ppGpp Hernandez V.J. J. Biol. 2001; PubMed D. Mol. PubMed PubMed Scholar). has been to to the active center by the and The presence of ppGpp the of the in that transcription stringent and is in the RNA polymerase have been which polymerase and initiation of RNA polymerase at ppGpp V.J. M. J. Mol. Biol. PubMed Scholar). This has been observed for other and as M. T. PubMed T. J. Biol. PubMed Scholar). is or is by its promoter is known for that stringent regulation of the promoter J. PubMed Scholar). This is termed the and has been to be for stringent regulation. is that other of in to the are in transcription is stringent or M. R. J. Scholar). in the have that ppGpp-dependent are by of their binary open the stable open complexes in presence of their M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed M. R. J. PubMed R.L. PubMed T. Gourse R.L. PubMed Scholar). the of the of open binary complexes is very and complexes have to at NTP formation of the ternary complexes be the step for transcription and at which step of the initiation ppGpp transcription this we have analyzed the effect of ppGpp on the steps to transcription initiation at P1 were and the promoter as We show that ppGpp the at the the binary open complexes to the ternary initiation The of the substrate in this that competition between the substrate and ppGpp for the is This is by almost identical inhibition characteristics ppGpp is by NTP substrate known to bind to the active of RNA polymerase. this we have to of the that are still with to the of the stringent This of regulation is by the effector molecule which transcription of in stable RNA The of this global changes in response to of environmental for which amino acid is the to the of this regulatory of bacterial we still are are and in is for that RNA polymerase is the for ppGpp Hernandez V.J. J. Biol. 2001; PubMed D. Mol. PubMed PubMed the are is known that the promoter is stringent or M. R. J. PubMed J. J. PubMed M. R. PubMed Scholar). and of transcription are by be effect of the ribosomal RNA the for stringent for the protein or amino acid are known to be at ppGpp of the recent that the of genes is by (2Barker M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: 689-702Crossref PubMed have been H.E. J. Biol. PubMed PubMed J. PubMed the of the stringent steps of the transcription cycle have been to be for regulation. formation of the open promoter complex has been as the step for the J. Biol. PubMed D. PubMed Scholar). in the closed complex by inhibition at promoter by of and promoter M. R. J. PubMed Scholar). The model is with the model, which suggests that RNA polymerase in with of the is triggered by bound ppGpp. RNA polymerase ppGpp stable RNA promoters, the with bound ppGpp J. Biol. PubMed Scholar). on the that stable RNA open for and stringent have been M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed M.M. Gourse R.L. J. 2001; PubMed Scholar). to ppGpp has effect on stringent stable RNA stringent regulation is as effect of RNA polymerase which results transcriptional shut-down of the stable RNA in and in vitro for transcriptional inhibition by ppGpp at the promoter Hernandez V.J. J. Biol. PubMed J. PubMed Scholar). the promoter stable open complexes that ppGpp-dependent regulation is to with open based on for stringent have been previously PubMed Ingraham J. J. J. Mol. Biol. PubMed Scholar). The and the that by itself the that is one for the stringent This is by the that ppGpp is to steps transcription initiation M. R. J. Biol. PubMed J. PubMed J. Biol. PubMed this we have in to which of the initiation are by ppGpp and to the molecular by which ppGpp exerts regulatory We the ribosomal RNA P1 the and which in their nucleotide for and for in the model, which that the NTP in the the of stringent T. Gourse R.L. PubMed Scholar). we the which stringent and which is known to be by ppGpp. with the we very inhibition in transcription the stable RNA were inhibited the conditions. the ppGpp inhibition of the and P1 promoter inhibition with for this is by the of the promoter P1 in and in the or the the that the major effect of ppGpp is or the formation of the ternary transcription initiation This be the inhibition is observed ppGpp is or at the formation of the ternary complex the initiation cycle initiation or to inhibition we show that the of initiating is for the of inhibition. NTP ppGpp-dependent inhibition We could that the the open complex is very in presence of ppGpp. We this as that the of ternary transcription complexes are dramatically by ppGpp with M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed we that the in of the ternary complexes in presence or of ppGpp is to the of inhibition we that the of ternary complex formation is reduced in presence of ppGpp. This effect be by an in the of the initiating these results are with previously published model, that ppGpp with the initiating NTP substrates R. J. Mol. Scholar). The model the that the for substrate to RNA polymerase active center is the by of the and M. PubMed Scholar). The of this nucleotide is to of one NTP at to ppGpp. is that at ppGpp of substrate to the active is This the for substrate binding are and the complexes are the for substrate are as is the for initiation complexes at stable RNA promoters, the competition Stringent be by the promoter which its and with RNA the of the initiating complex and the for the first substrate The effector molecule which to the stringent competition to the ppGpp with the substrate for access to the active center of RNA polymerase. or initiation complexes are are in an to previously published J. Biol. PubMed that ppGpp the of RNA polymerase by of the transcription The by that ppGpp-dependent inhibition to be with the triphosphate are for this the of ppGpp-dependent inhibition on be that on ternary initiation complex formation the promoter is The were at with transcription complexes at the and were is the to the complex with on The observed inhibition be the of one inhibition and inhibition be complex substrate competition is in the of ppGpp-dependent inhibition by the effector molecule ppGpp with nucleotide We and that are known to be as and are to the active center of RNA polymerase. be seen in and these inhibited transcription of the stable RNA almost the characteristics of inhibition and were at the as ppGpp. in the promoter for the or at the step to formation of the ternary initiation the on the of the as observed with the reduced the at which the ternary complexes are We that these are an we these as that substrate competition major in the of stringent is the that substrates other of ppGpp-dependent inhibition. substrates with ppGpp binding and in the as be the for ppGpp. recent in initiation the at the that inhibition Hernandez V.J. J. Biol. PubMed Scholar). the of this the that ppGpp formation of the first and this step is by the that the effect of ppGpp is in the presence of the The as to this the model and to the of inhibition or with are is for the regulation of NTP access to the active of RNA and the for of NTP substrates has been in the that is for the have been the RNA the by and or are to for RNA polymerase The with the at the active of RNA polymerase. The the and the for the active This termed by the of the and the of substrate Mol. PubMed of competition on RNA polymerase the complex be changes of the transcription are in of the that RNA polymerase be by as in the of the the of the is by in the molecule binding to stringent or the with an effector molecule ppGpp the of the could of the stringent which to be substrate have that ppGpp the of with RNA polymerase M. T. PubMed T. J. Biol. PubMed T. J. Biol. PubMed Scholar). this ppGpp could be an effector that at the and which the complex of the that is known to with the PubMed PubMed Scholar). this we have to of the that are still with to the of the stringent This of regulation is by the effector molecule which transcription of in stable RNA The of this global changes in response to of environmental for which amino acid is the to the of this regulatory of bacterial we still are are and in is for that RNA polymerase is the for ppGpp Hernandez V.J. J. Biol. 2001; PubMed D. Mol. PubMed PubMed the are is known that the promoter is stringent or M. R. J. PubMed J. J. PubMed M. R. PubMed Scholar). and of transcription are by be effect of the ribosomal RNA the for stringent for the protein or amino acid are known to be at ppGpp of the recent that the of genes is by (2Barker M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: 689-702Crossref PubMed have been H.E. J. Biol. PubMed PubMed J. PubMed Scholar). the of the stringent steps of the transcription cycle have been to be for regulation. formation of the open promoter complex has been as the step for the J. Biol. PubMed D. PubMed Scholar). in the closed complex by inhibition at promoter by of and promoter M. R. J. PubMed Scholar). The model is with the model, which suggests that RNA polymerase in with of the is triggered by bound ppGpp. RNA polymerase ppGpp stable RNA promoters, the with bound ppGpp J. Biol. PubMed Scholar). on the that stable RNA open for and stringent have been M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed M.M. Gourse R.L. J. 2001; PubMed Scholar). to ppGpp has effect on stringent stable RNA stringent regulation is as effect of RNA polymerase which results transcriptional shut-down of the stable RNA in and in vitro for transcriptional inhibition by ppGpp at the promoter Hernandez V.J. J. Biol. PubMed J. PubMed Scholar). the promoter stable open complexes that ppGpp-dependent regulation is to with open based on for stringent have been previously PubMed Ingraham J. J. J. Mol. Biol. PubMed Scholar). The and the that by itself the that is one for the stringent This is by the that ppGpp is to steps transcription initiation M. R. J. Biol. PubMed J. PubMed J. Biol. PubMed Scholar). this we have in to which of the initiation are by ppGpp and to the molecular by which ppGpp exerts regulatory We the ribosomal RNA P1 the and which in their nucleotide for and for in the model, which that the NTP in the the of stringent T. Gourse R.L. PubMed Scholar). we the which stringent and which is known to be by ppGpp. with the we very inhibition in transcription the stable RNA were inhibited the conditions. the ppGpp inhibition of the and P1 promoter inhibition with for this is by the of the promoter P1 in and in the or the the that the major effect of ppGpp is or the formation of the ternary transcription initiation This be the inhibition is observed ppGpp is or at the formation of the ternary complex the initiation cycle initiation or to inhibition we show that the of initiating is for the of inhibition. NTP ppGpp-dependent inhibition We could that the the open complex is very in presence of ppGpp. We this as that the of ternary transcription complexes are dramatically by ppGpp with M.M. Gaal T. Gourse R.L. J. Mol. Biol. 2001; 305: PubMed we that the in of the ternary complexes in presence or of ppGpp is to the of inhibition we that the of ternary complex formation is reduced in presence of ppGpp. This effect be by an in the of the initiating these results are with previously published model, that ppGpp with the initiating NTP substrates R. J. Mol. Scholar). The model the that the for substrate to RNA polymerase active center is the by of the and M. PubMed Scholar). The of this nucleotide is to of one NTP at to ppGpp. is that at ppGpp of substrate to the active is This the for substrate binding are and the complexes are the for substrate are as is the for initiation complexes at stable RNA promoters, the competition Stringent be by the promoter which its and with RNA the of the initiating complex and the for the first substrate The effector molecule which to the stringent competition to the ppGpp with the substrate for access to the active center of RNA polymerase. or initiation complexes are are in an to previously published J. Biol. PubMed that ppGpp the of RNA polymerase by of the transcription The by that ppGpp-dependent inhibition to be with the triphosphate are for this the of ppGpp-dependent inhibition on be that on ternary initiation complex formation the promoter is The were at with transcription complexes at the and were is the to the complex with on The observed inhibition be the of one inhibition and inhibition be complex substrate competition is in the of ppGpp-dependent inhibition by the effector molecule ppGpp with nucleotide We and that are known to be as and are to the active center of RNA polymerase. be seen in and these inhibited transcription of the stable RNA almost the characteristics of inhibition and were at the as ppGpp. in the promoter for the or at the step to formation of the ternary initiation the on the of the as observed with the reduced the at which the ternary complexes are We that these are an we these as that substrate competition major in the of stringent is the that substrates other of ppGpp-dependent inhibition. substrates with ppGpp binding and in the as be the for ppGpp. recent in initiation the at the that inhibition Hernandez V.J. J. Biol. PubMed Scholar). the of this the that ppGpp formation of the first and this step is by the that the effect of ppGpp is in the presence of the The as to this the model and to the of inhibition or with are is for the regulation of NTP access to the active of RNA and the for of NTP substrates has been in the that is for the have been the RNA the by and or are to for RNA polymerase The with the at the active of RNA polymerase. The the and the for the active This termed by the of the and the of substrate Mol. PubMed Scholar). The of competition on RNA polymerase the complex be changes of the transcription are in of the that RNA polymerase be by as in the of the the of the is by in the molecule binding to stringent or the with an effector molecule ppGpp the of the could of the stringent which to be substrate have that ppGpp the of with RNA polymerase M. T. PubMed T. J. Biol. PubMed T. J. Biol. PubMed Scholar). this ppGpp could be an effector that at the and which the complex of the that is known to with the PubMed PubMed Scholar).