antitermination of transcription of catabolic operons

MicroReview

Antitermination of transcription of catabolic operons

Blanka RutbergDepartment of Microbiology, Lund University, Solvegatan21, S-22362 Lund, Sweden.

Summary

Antitermination of transcription mediated by proteinsinteracting with mRNA sequences is described fornine operons/regulons. Eight of the systems are cata-bolic, while the ninth, the Klebsiella pneumoniae nasregulon, is involved in the assimilation of nitrate andnitrite. Six of the catabolic operons/regulons are foundin Bacillus subtilis , one is found in Escherichia coli ,and one in Pseudomonas aeruginosa . The antitermi-nation system of five of the operons/regulons ( E.coli blg , and sacPA , sacB , bgl , and lic from B. subtilis )are assigned to the bgl–sac family on the basis ofextensive similarities with regard to antiterminatorproteins and the sequences of the antiterminators.Other members of the bgl–sac family are the arboperon of Erwinia chrysanthemi and a presumed bgloperon of Lactococcus lactis . The antiterminationsystems of the other four operons/regulons ( B. subti-lis glp , B. subtilis hut , P. aeruginosa ami , and K. pneu-moniae nas ) seem to be unrelated both to the bgl–sacfamily and to each other. The antiterminator protein ofthe B. subtilis glp regulon has been found not only tocause antitermination but also to stabilize the result-ant mRNA and to mediate glucose repression. If otherantiterminator proteins, and antitermination factors,also prove to have additional functions, it will broadenthe impact of antitermination as a means of control-ling gene expression.

Introduction

Expression of bacterial genes is most often regulated atthe transcriptional level. The number of transcripts canbe controlled by the frequency of transcription initiation,and the length of transcripts can be varied by prematuretranscription termination. Rho-independent transcriptionterminators located in the leader or an intercistronic regionof an operon can result in termination of transcription and

truncated transcripts. Antitermination which gives rise tofull-length transcripts can be achieved in two principalways: (i) RNA polymerase can be modified so that itreads through transcription terminators, as is found inthe phage l lytic developmental pathway (Greenblatt etal., 1993); or (ii) formation of the terminator can be pre-vented by the interaction (or lack of interaction) of a factorwith mRNA as described below. Some authors use theterm attenuation for the second type of antitermination(Landick and Turnbough, 1992) but, in general, attenua-tion seems best reserved for the mechanism that is repre-sented by the Escherichia coli biosynthetic trp operon.Here attenuation is effected by ribosomes which stall inthe absence of charged tRNATrp and then prevent forma-tion of a terminator (Yanofsky and Crawford, 1987).

Antitermination resulting from factors controlling the for-mation of a terminator occurs in many operons and in avariety of different ways. The Bacillus subtilis trp operonis negatively regulated by a protein called TRAP (trpattenuator protein). In the presence of tryptophan, the11-subunit TRAP binds to several (G/U)AG repeats inthe trp leader mRNA (Babitzke et al., 1996) and inducesformation of a terminator. In the absence of tryptophan,TRAP is unable to bind and an antiterminator forms allow-ing operon transcription (Gollnick, 1994). The antitermina-tor is a stem–loop structure which overlaps the terminatorby four bp and prevents its formation. The B. subtilis purand pyr operons seem to be regulated by a similar mech-anism (Zalkin and Ebbole, 1988; Turner et al., 1994).

Many aminoacyl-tRNA-synthetase and amino acid bio-synthesis genes in several Gram-positive genera appearto be regulated in response to the availability of the cog-nate amino acid using uncharged tRNA as the positiveregulator (Henkin, 1994). Genetic analysis of the B. sub-tilis tyrS gene has shown that uncharged tRNATyr interactswith the tyrS leader mRNA and promotes formation of anantiterminator which results in tyrS expression.

Antitermination in the catabolic operons described belowis substrate induced and is effected by antiterminator pro-teins that have been shown, or are proposed, to bind to themRNA. Six of the systems are found in the Gram-positive,low-GC-content bacterium B. subtilis, and three are foundin the Gram-negative bacteria E. coli, Pseudomonas aeru-ginosa and Klebsiella pneumoniae, respectively. The K.pneumoniae nas regulon is involved in nitrate assimilationand is thus not a catabolic regulon but it is regulated by an

Molecular Microbiology (1997) 23(3), 413–421

Q 1997 Blackwell Science Ltd

Received 17 September, 1996; revised 5 November, 1996; accepted8 November, 1996. E-mail [email protected]; Tel. (46)2228629; Fax (46) 157839.

m

antitermination mechanism. The E. coli system and four ofthe B. subtilis systems can be grouped together on thebasis of extensive similarities, while each of the remainingfour systems seems to be of a different origin.

In the literature, there is some confusion about the wordantiterminator. It is used to denote both a protein thatcauses antitermination and an mRNA sequence with thepotential to form a secondary structure which interfereswith the formation of a terminator. In the following, antiter-minator will be used in the second sense to denote anmRNA sequence, while a protein causing antiterminationwill be referred to as an antiterminator protein.

The bgl operon of E. coli

The bgl operon of E. coli (Fig. 1) contains three genesinvolved in the utilization of aromatic b-glucosides: bglG,which encodes an antiterminator protein; bglF, whichencodes a transporter of b-glucosides and belongs tothe phosphotransferase (PTS) family; and bglB, the struc-tural gene that encodes a phospho-b-glucosidase. bglG isflanked by sequences defining rho-independent transcrip-tion terminators (Bramley and Kornberg, 1987; Maha-devan et al., 1987; Schnetz et al., 1987). BglG binds tothe leader mRNA just upstream of the first terminator(Houman et al., 1990) and allows transcription read-through (Mahadevan and Wright, 1987; Schnetz andRak, 1988). The 32-nucleotide target sequence of BglGextends into the terminator and has the potential to forman alternative stem–loop structure which is less stablethan the terminator. This sequence has been namedRAT (ribonucleic antiterminator; Aymerich and Steinmetz,1992). Binding of BglG is thought to stabilize the RAT andpreclude formation of the terminator. Unpublished experi-ments indicate that BglG binds to an almost identical RATpartially overlapping the second terminator which liesbetween bglG and bglF (Amster-Choder and Wright,1993). In the absence of b-glucosides, BglG is phosphory-lated by BglF and is inactive in antitermination. BglF ishomologous to enzyme IIglucose (EIIGlc) of the PTS andis also called EIIbgl. Phosphate is transferred fromphosphoenolpyruvate via intermediates to BglF and thento b-glucosides or BglG (Amster-Choder and Wright,

1990; Schnetz and Rak, 1990). In its phosphorylatedform, BglG is monomeric and is unable to bind to theRAT, while non-phosphorylated BglG dimerizes andbinds to the RAT (Amster-Choder and Wright, 1992).

Erwinia chrysanthemi, a relative of E. coli, contains anoperon (arb ) which is homologous to the E. coli bgl operon(El Hassouni et al., 1992). The Gram-positive, low-GCbacterium Lactococcus lactis also appears to contain abgl operon homologous to that of E. coli, although it hasonly been partially characterized (Bardowski et al., 1994).

The sac regulons of B. subtilis

The B. subtilis sac regulons are involved in the utilization ofsucrose. sacA encodes an intracellular phosphosucrase(Fouet et al., 1986) and sacB encodes an extracellularlevansucrase (Steinmetz et al., 1985). sacP encodes aPTS component, enzyme IIsucrose (EIIScr), which mediatestransport of sucrose (Fouet et al., 1987). The sacPA andsacB operons (Fig. 2) are induced by the antiterminatorproteins SacT and SacY, respectively, in the presence ofsucrose (Debarbouille et al., 1990; Crutz et al., 1990).Full induction of sacPA and sacB is reached at 1 mM and30 mM sucrose, respectively. sacY is part of the sacXYoperon and sacX encodes a protein which is homologousto SacP (56% identity) and to several other EIIScr

(Zukowski et al., 1990). The leader transcripts of thesacPA and sacB genes each contain a RAT-terminatorsequence (Steinmetz et al., 1985; Shimotsu and Henner,1986; Debarbouille et al., 1990). Recently, gel retardationexperiments have shown that SacT and SacY bind tosacPA leader mRNA (Arnaud et al., 1996). SacT andSacY are 48% identical to each other and 35% identicalto BglG of E. coli. There is cross-talk between the sacoperons under certain conditions, i.e. SacT can stimulatesacB expression and SacY can stimulate sacPA expres-sion (Steinmetz et al., 1989). SacY can also substitutefor BglG and mediate expression of the E. coli bgl operon;however, SacY is not regulated by E. coli BglF (Amster-Choder and Wright, 1993). The RATs of sacPA and

Q 1997 Blackwell Science Ltd, Molecular Microbiology, 23, 413–421

Fig. 1. Schematic representation of the E. coli bgl operon and itsproposed transcripts (arrows). Stem–loops symbolize transcriptionterminators. The black box denotes a gene encoding anantiterminator protein. The dotted box denotes a gene encoding aPTS component that phosphorylates and inactivates theantiterminator protein.

Fig. 2. Schematic representation of (A) the B. subtilis sacPAregulon and (B) the B. subtilis sacB regulon and their proposedtranscripts (arrows). Stem–loops symbolize transcriptionterminators. The dotted stem–loop symbolizes a putativeterminator.

414 B. Rutberg

sacB differ from each other at three positions. In vitromutagenesis has shown that these differences are respon-sible for the specific induction of sacPA by SacT and sacBby SacY (Aymerich and Steinmetz, 1992).

The activity of SacY is regulated by SacX, probably byphosphorylation/dephosphorylation (Crutz et al., 1990),analogous to the regulation of BglG by BglF in E. coli. Incontrast to BglF, however, SacX does not contain anEIIA domain (Saier et al., 1993). It therefore has onlyone phosphorylation site and it is not clear which PTS ele-ment is used to phosphorylate SacX. The activity of SacTis both positively and negatively regulated by the PTS. Ithas been proposed that SacT has two sites for phosphory-lation. One is an activation site which is phosphorylated bythe general components of the PTS (EI and HPr). The sec-ond is an inactivation site which is phosphorylated in theabsence of sucrose and dephosphorylated in its presence.To be active in antitermination, SacT has to be phosphory-lated at its activation site and dephosphorylated at its inac-tivation site (Arnaud et al., 1992). It seems plausible thatactive SacT, like active BglG, is a dimer and that phos-phorylation by the general PTS components facilitatesdimerization. However, it appears that non-phosphory-lated SacT binds to mRNA to the same extent as SacTthat has been phosphorylated by HPr (Arnaud et al.,1996). The explanation could be that high concentrationsof SacT allow dimerization independently of the PTS. Thishypothesis is supported by experiments on LicT, whichis another antiterminator protein of the bgl–sac family(see below). It is not known which intermediate is involvedin phosphorylation of the inactivation site of SacT. SacP(EIIScr), which like SacX does not have an EIIA domain,is not required for SacT activity.

The sacXY operon is also sucrose inducible. At low con-centrations of sucrose, induction is mediated by SacT,while at higher concentrations it is mediated by bothSacT and SacY. The sacXY leader transcript contains aRAT-like sequence and, at a distance of 100 bp, a potentialterminator sequence. In spite of this distance, formation ofthe RAT and the potential terminator might be mutuallyexclusive according to RNA modelling (Tortosa and LeCoq, 1995).

The lic and bgl operons of B. subtilis

The lic and bgl operons of B. subtilis (Fig. 3) are bothcontrolled by the antiterminator protein LicT. LicT is verysimilar to BglG of E. coli and SacT and SacY of B.

subtilis (Schnetz et al., 1996). It causes antitermination ata RAT-terminator sequence situated between licT andlicS. licS encodes an extracellular b-glucanase and anti-termination is induced in the presence of the b-glucanlichenan, tetrasaccharides which are degradation pro-ducts of lichenan or the b-glucoside salicin.

LicT also causes antitermination at a RAT-terminatorsequence in the leader of the bglPH operon (Kruger andHecker, 1995; Le Coq et al., 1995). bglP encodes EIIbgl,a b-glucoside-specific enzyme of the PTS, and bglHencodes a phospho-b-glucosidase. In contrast with licS,antitermination of bglPH is induced by b-glucosides butnot by lichenan. LicT and BglP are tied in a regulatorycircuit. Expression of bglP depends on antiterminationcaused by LicT in the presence of b-glucosides. LicT isinactivated by BglP, possibly by phosphorylation via thePTS analogous to the regulation of BglG by BglF (EIIbgl)in E. coli. Through its action on LicT, BglP negatively reg-ulates its own synthesis. LicT is also positively regulatedby the general components of the PTS and it is possiblethat LicT is active as a dimer and that phosphorylationby the PTS enhances dimerization. The finding that anincreased number of LicT copies overcomes the depen-dence of bglP expression on the PTS supports this model(Kruger and Hecker, 1995).

There is some cross-talk between LicT and the otherantiterminator proteins of the bgl–sac family. LicT causesantitermination in the bgl operon of E. coli and E. coli BglGcauses antitermination in the licTS operon (Schnetz et al.,1996). LicT may also cause antitermination in the sac oper-ons, because a strain from which sacT and sacY had beendeleted regained the ability to use sucrose through inacti-vation of bglP, but lost this ability if licT was also inacti-vated (Steinmetz and Richter, 1994; Le Coq et al., 1995).On the other hand, SacT and SacY do not appear to induceantitermination in the bglPH operon (Kruger and Hecker,1995). Although LicT induces antitermination of both licSand bglPH, there is a difference in the regulation of antiter-mination activity in the two operons, because antitermina-tion of licS but not of bglPH can be induced by lichenan.

The glp regulon of B. subtilis

The B. subtilis glp regulon (Fig. 4) is involved in the uptakeand catabolism of glycerol and glycerol-3-phosphate(G3P). It contains four operons: glpP, glpFK, glpD andglpTQ (Lindgren and Rutberg, 1974; Holmberg et al.,1990; Beijer et al., 1993; Nilsson et al., 1994). glpP encodes


Fig. 3. Schematic representation of (A) the B.subtilis lic operon and its transcripts (arrows)and (B) the B. subtilis bgl operon and itsproposed transcripts (arrows). Stem–loopssymbolize transcription terminators.

Antitermination of catabolic operons 415

an antiterminator protein, glpF encodes a glycerol uptakefacilitator, glpK a glycerol kinase, glpD a G3P dehydrogen-ase, glpT a G3P permease and glpQ a glycerophospho-diester phosphodiesterase. The leaders of glpFK, glpDand glpTQ contain sequences which define transcriptionterminators. The sequences are very similar to eachother in the region that corresponds to the lower part ofthe terminator stem and there are conserved regions justupstream of the terminator sequences. However, RNAmodelling does not disclose any potential antiterminatorsequences overlapping the terminator sequences. GlpPinduces expression of the glp operons in the presence ofG3P. The sequence of GlpP reveals a possible phos-phate-binding site near the C-terminus of the protein(Bork et al., 1995), but apart from that, GlpP is not similarto any protein found in the data banks. For the glpD operonit has been shown that full-length transcripts are obtainedunder inducing conditions, while under non-inducing con-ditions, the transcripts terminate at nucleotides in thedownstream stem of the terminator (Holmberg and Rut-berg, 1991; 1992). Point mutations and deletions within theglpD terminator sequence lead to constitutive, GlpP- andG3P-independent expression of glpD (Glatz et al., 1996).They also result in glucose-repression-insensitive expres-sion of glpD.

In addition to the constitutive glpD phenotype, mutationsin the loop part of the glpD terminator sequence have beenshown to cause instability of glpD mRNA (Glatz et al.,1996). This is clearly seen at 458C where the half-life ofmutant glpD mRNA is less than 0.3 min while the half-lifeof wild-type glpD mRNA is approx. 2.5 min. Mutations inthe stem part of the glpD terminator sequence do notcause this effect. The instability of the mutant glpD mRNAis seen only in GlpP-deficient strains. When the wild-typeglpP gene is introduced into these strains the glpD mRNAis stabilized. Introduction of glpP also restores glucoserepression indicating that it is mediated by the GlpP protein(Glatz et al., 1996).

The hut operon of B. subtilis

The B. subtilis hut operon (Fig. 5) encodes enzymes forthe catabolism of L-histidine (Kimhi and Magasanik, 1970).It is regulated by an antiterminator protein, HutP, whichis not similar to other antiterminator proteins (Oda et al.,

1988; Wray and Fisher, 1994). The operon contains apotential transcription terminator but, as yet, no antitermi-nator sequence has been identified.

The ami operon of P. aeruginosa

The amidase (ami) operon of P. aeruginosa (Fig. 6) con-tains five genes that are involved in the utilization ofshort-chain aliphatic amides, such as acetamide (Bram-mar and Clarke, 1964). amiE encodes an amidase, amiBand amiS are thought to be part of an amide transport sys-tem, amiR encodes an antiterminator protein, and amiCencodes a protein which regulates AmiR activity (Bram-mar et al., 1987; Cousens et al., 1987; Drew and Lowe,1989; Lowe et al., 1989; Wilson et al., 1993). Upstreamof amiE there is a strong promoter followed by a transcrip-tion terminator sequence. Northern analysis has showntwo classes of transcripts under inducing conditions: oneis the result of termination between amiE and amiB, andthe other is a full-length transcript representing the wholeoperon (Wilson and Drew, 1995). A third class of transcriptmay be initiated at a promoter between amiB and amiC.Just upstream of the first terminator sequence there is asequence showing some similarity to the RAT sequen-ces. In vivo titration experiments demonstrate that the anti-terminator protein, AmiR, interacts with the amiE leadermRNA (Wilson et al., 1996). AmiR activity is negativelyregulated by AmiC and an AmiC–AmiR stable dimer–dimer complex has been isolated in the presence of theanti-inducing ligand butyramide. Acetamide, the inducer,binds to AmiC and stimulates antitermination by causingdissociation of the AmiC–AmiR complex (Wilson et al.,1996). Sequence and structural determinations haveidentified AmiC as a member of the structural family ofperiplasmic binding proteins although it is found in thecytoplasm (Pearl et al., 1994). It appears in this casethat a simple steric hindrance mechanism is responsible


Fig. 4. Schematic representation of the B. subtilis glp regulon and its transcripts (arrows). The dotted arrows indicate proposed transcripts.Stem–loops symbolize transcription terminators.

Fig. 5. Schematic representation of the B. subtilis hut operon andits proposed transcripts (arrows). The stem–loop symbolizes atranscription terminator.

416 B. Rutberg

for the regulation of antitermination activity which onlyoccurs at the upstream terminator.

The nas regulon of K. pneumoniae

K. pneumoniae can use nitrate and nitrite as sole nitrogensources during aerobic growth. The proteins required fornitrate assimilation are encoded by the nasFEDCBA oper-on (Fig. 7), also termed the nasF operon (Lin et al., 1993;1994). This non-catabolic operon is presented here becauseit seems to be regulated in much the same way as the cata-bolic operons described above. nasF, nasE and nasDencode what appears to be a nitrate/nitrite uptake system,nasC and nasA encode an assimilatory nitrate reductase,and nasB encodes an assimilatory nitrite reductase.Upstream of nasF lies nasR which encodes an antitermi-nator protein (Goldman et al., 1994). In the presence ofnitrate or nitrite, NasR induces expression of the nasFoperon, presumably by allowing transcription readthroughat a terminator in the leader of the nasF operon (Lin andStewart, 1996). Upstream of and overlapping the termina-tor there is a possible antiterminator that is not related tothe RATs. However, the antiterminator protein NasRshares C-terminal sequence similarity with AmiR.

Discussion

Table 1 summarizes some of the characteristics of theantitermination system that have been described. Exten-sive similarities are seen for the components of antitermi-nation in the bgl–sac operons (E. coli bgl, B. subtilis sac,B. subtilis bgl and B. subtilis lic). The arb operon of E. chry-santhemi, which appears closely related to the E. coli bgloperon, should be added to this group and presumablythere is a similarly regulated bgl operon in Lactococcus lac-tis which should also be included. The antiterminator pro-teins of the bgl–sac family are of similar sizes, showsequence homology, and exhibit some cross-talk. A com-parison of the amino acid sequences of the bgl–sac anti-terminator proteins is found in the article by Schnetz et

al. (1996). The antiterminator protein BglG of E. coli hasbeen shown to bind to an mRNA sequence that couldform a stem–loop structure, a RAT. Formation of theRAT would preclude formation of the terminator. Bindingof the antiterminator proteins SacT and SacY to sacPAleader mRNA has recently been shown. The bgl–sacoperons display very similar RAT-terminator sequences.The exception is the sacXY operon where the RATsequence is separated from the probable terminatorsequence by 100 bp. BglG is phosphorylated by the PTScomponent BglF (EIIbgl) that mediates transport of theinducing b-glucosides. Phosphorylated BglG is mono-meric and inactive in antitermination. In the presence ofb-glucosides, BglG is dephosphorylated which leads toits dimerization and activation. SacY appears to behavesimilarly, and the PTS component involved in this case isSacX (EIIScr). It is likely that SacT and LicT also dimerizeto be active but in their case the general components ofthe PTS might be responsible for the phosphorylationthat leads to activation. SacT is also negatively affectedby the PTS and it has been proposed that there are twophosphorylation sites on SacT: one activation site andone inactivation site.

The striking similarities between the antiterminationsystems of bacteria that are, in evolutionary terms, farapart, point to horizontal gene transfer. The antitermina-tion systems of the four operons/regulons in the lowerhalf of Table 1 (B. subtilis glp, B. subtilis hut, P. aeruginosaami and K. pneumoniae nas) do not resemble the bgl–sacsystems, nor do they appear to be related to each other.However, the amiE leader contains a sequence that issimilar in part to the RAT sequences, and the antitermina-tor proteins AmiR and NasF show sequence similarity attheir C-terminal ends. These antiterminator proteins areactivated in the presence of different types of compounds.GlpP, HutP and NasR may interact directly with theirrespective inducers, glycerol-3-phosphate, histidine andnitrate/nitrite, respectively; AmiR is negatively regulatedby the interaction with another protein, AmiC, to whichinducing amides bind.

No apparent antiterminator sequences have yet beendescribed in the B. subtilis glp and hut operons and theAmiR recognition sequence upstream of the first termina-tor in the P. aeruginosa ami operon is unable to form analternative stem–loop structure. Is antitermination in theseoperons mechanistically different from antitermination inthe bgl–sac operons? One could, for example, picturethe antiterminator protein simply binding close to theterminator region of the nascent mRNA and preventing


Fig. 6. Schematic representation of the P. aeruginosa ami operonand its transcripts (arrows). The dotted arrows indicate proposedtranscripts. The stem–loop symbolizes a transcription terminator.

Fig. 7. Schematic representation of the K.pneumoniae nas operon and its proposedtranscripts (arrows). The stem–loopsymbolizes a transcription terminator.



Tab

le1.

Som

ech

arac

teri

stic

sof

antit

erm

inat

ion

inei

ght

cata

bolic

oper

ons/

regu

lons

and

the

K.

pneu

mon

iae

nas

oper

on.

Ope

ron

/reg

ulon

Ant

iterm

inat

orpr

otei

nIn

duce

rof

antit

erm

inat

ion

DG

0of

term

inat

or(s

)A

ntite

rmin

ator

sequ

ence

(s)

Rem

arks

E.

coli

bgl

Bgl

G;

278

aaa,

32kD

a;au

tore

gula

ted

b-g

luco

side

s,e.

g.sa

licin

¹88

kJm

ol¹

1;

¹10

9kJ

mol

¹1;

flank

ing

bglG

2R

AT

sbB

glG

isde

phos

phor

ylat

ed/a

ctiv

ated

byB

glF

;ac

tive

Bgl

Gis

adi

mer

B.

subt

ilis

sacP

AS

acT

;27

7aa

,32

kDa;

sim

ilar

toB

glG

Suc

rose

,1

mM

¹13

0kJ

mol

¹1

1R

AT

Sac

Tis

activ

ated

byth

ege

nera

lcom

pone

nts

ofth

eP

TS

B.

subt

ilis

sacB

Sac

Y;

280

aa,

32kD

a;si

mila

rto

Bgl

GS

ucro

se,

30m

Msa

cB,¹

166

kJm

ol¹

1;

sacX

Y,¹

49kJ

mol

¹1

2R

AT

sS

acY

is(p

roba

bly)

deph

osph

oryl

ated

/act

ivat

edby

Sac

X;

upst

ream

ofsa

cXY

,a

RA

Tse

quen

cean

da

poss

ible

term

inat

orse

quen

cear

ese

para

ted

by10

0bp

Bsu

btili

sbg

lLi

cT;

277

aa,

32kD

a;si

mila

rto

Bgl

Gb

-glu

cosi

des,

e.g.

salic

in¹

126

kJm

ol¹

11

RA

TLi

cTis

activ

ated

byth

ege

nera

lcom

pone

nts

ofth

eP

TS

B.

subt

ilis

licT

etra

sacc

hari

des

from

liche

nan,

salic

in¹

79kJ

mol

¹1

1R

AT

B.

subt

ilis

glp

Glp

P;

192

aa,

22kD

aG

lyce

rol-3

-pho

spha

te(G

3P)

glpF

K,¹

91kJ

mol

¹1;

glpD

,¹

147

kJm

ol¹

1;

glpT

Q,¹

142

kJm

ol¹

1

Non

eid

entif

ied

Glp

Pst

abili

zes

glpD

mR

NA

and

med

iate

sgl

ucos

ere

pres

sion

ofgl

pD

B.

subt

ilis

hut

Hut

P;

151

aa,

17kD

aH

istid

ine

¹11

5kJ

mol

¹1

Non

eid

entif

ied

P.

aeru

gino

saam

iA

miR

;19

6aa

,22

kDa

Sho

rt-c

hain

alip

hatic

amid

es¹

126

kJm

ol¹

1,

2/3

ofa

RA

TA

miR

isne

gativ

ely

regu

late

dby

Am

iC;

Am

iCis

inac

tivat

edby

amid

es

K.

pneu

mon

iae

nas

Nas

R;

393

aa,

44kD

aN

itrat

e/n

itrite

¹85

kJm

ol¹

11

sequ

ence

not

rela

ted

toth

eR

AT

sN

ota

cata

bolic

oper

on

a.aa

,am

ino

acid

(s).

b.R

AT

(rib

onuc

leic

antit

erm

inat

or)

isth

em

RN

Ata

rget

sequ

ence

(,30

nucl

eotid

eslo

ng)

for

antit

erm

inat

orpr

otei

nsof

the

bgl–

sac

fam

ily.

418 B. Rutberg

terminator stem–loop formation. Alternatively, antitermi-nation in these operons may depend on the formation ofantiterminators which will only be discerned as we gain abetter understanding of RNA folding (Westhof and Michel,1994).

The antiterminator protein GlpP has two additional func-tions: it can stabilize glpD mRNA and it mediates glucoserepression of glpD. Kruger et al. (1996) have recently sug-gested that a glucose repression mechanism which isindependent of the repressor CcpA and its target sequ-ence CRE (catabolite repression element) (Saier et al.,1996) is mediated via LicT. Except for sacB, the catabolicoperons discussed here are all sensitive to cataboliterepression by glucose or, in the case of P. aeruginosa,succinate (Reynolds et al., 1986; Lepesant et al., 1976;Kruger and Hecker, 1995; Kruger et al., 1993; Chasinand Magasanik, 1968; Smyth and Clarke, 1975). It is aninteresting possibility that antiterminator proteins mightplay a role in catabolite repression.

It has been proposed that in addition to causing antiter-mination, SacT and SacY also stabilize the antiterminatedtranscripts (Crutz and Steinmetz, 1992). Antiterminatorproteins of catabolic operons are proposed to bind to spe-cific mRNAs, and BglG, SacT and SacY have been shownso to do. Could it be that they also stabilize the resultanttranscripts? Could it even be that factors interacting withmRNA and causing antitermination in anabolic operonsalso stabilize the mRNA? To answer these questions onewould need experimental conditions which allow themRNA-stabilizing effect of antitermination factors to beseparated from their antiterminating activity.

Currently, relatively few cases of antitermination regula-tion of catabolic operons are known. Most probably manymore cases of control of gene activity in bacteria by termi-nation/antitermination of transcription will be described inthe future for catabolic and other operons. One recentexample shows that 70–80% of the transcripts of a pepti-dase gene in Streptococcus pyogenes are prematurelyterminated at an inverted repeat preceding the gene (Prit-chard and Cleary, 1996). However, a role for this repeat incontrolling gene activity has not yet been demonstrated. Ifantitermination factors can also mediate stabilization ofthe mRNA which they give rise to, as shown for the B.subtilis GlpP protein, this would add to the sophisticationand usefulness of termination/antitermination as a meansof controlling gene expression.

Antiterminator proteins of several of the systems dis-cussed here seem unrelated by sequence comparisons,which suggests that this type of control may have evolvedon several independent occasions. However, the closelyrelated bgl–sac systems are represented both in E. coliand B. subtilis, strongly suggesting the occurrence ofhorizontal gene transfer. We know rather little about theregulatory mechanisms of the antitermination systems

described in this review and we know even less aboutthe nature of the interactions between the antiterminatorproteins and their mRNAs. Also, the role of antiterminatorproteins in global regulatory circuits in carbon catabolismsuch as glucose repression in B. subtilis and other eubac-teria needs to be further explored. Clearly, studies oftermination/antitermination systems, such as those des-cribed here and others yet to be discovered, will give newinformation on problems of general biological significance.

Acknowledgements

I thank Per Johansson for searching the literature on antiter-mination in databases, and Mikael Matsson for preparing thefigures. I am grateful to Lars Rutberg for critically reading themanuscript and to Robert Drew for communicating data inpress. Work on the B. subtilis glp regulon was supported bygrants from the Swedish Medical Research Council, the Emiland Wera Cornell Foundation, and the Crafoord Foundation.

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