Protein signaling via type III secretion pathways in phytopathogenic bacteria

109

Protein signaling via type III secretion phytopathogenic bacteria Mary Beth Mudgett* Progress

in the genetic

and Brian J Staskawiczt

and biochemical

hrp-encoded

type III secretion

mechanisms

by which phytopathogenic

The suggestion

dissection

of the

pathway has revealed new

by’ plants cells has fundamentally

the way in which plant-bacterial

interactions

changed

are now being

viewed.

Addresses Department

of Plant and Microbial Biology, 111 Koshland Hall,

University of California, Berkeley, CA 94720-3102,

USA

*e-mail: [email protected] +,-mail: [email protected]

Current Opinion in Microbiology

1998,

1 :109-l

14

http://biomednet.com/elecref/1369527400100109 0 Current

Biology Ltd ISSN

1369-5274

Abbreviations HR

hypersensitive

Pv NLS

pathovar nuclear localization signals

response

Introduction Plants are continually exposed to a number of potentially pathogenic bacteria. Phytopathogenic bacteria, in general, are intercellular pathogens, invading and multiplying only in the spaces between plant cells. By evolving precise defense pathways, plants actively recognize invading bacteria to impede their ingress and colonization. The phenotype of plant disease resistance is the rapid induction of the hypersensitive programmed cell death pathway. In many plant-pathogen interactions, the hypersensitive response (HR) is associated with the production of reactive oxygen intermediates, the alteration of ion fluxes, the oxidative cross-linking of cell wall structural proteins, and the synthesis of antimicrobial compounds, including phytoalexins and pathogenesis-related proteins [1,2]. The mechanism(s) for pathogen inhibition are not known; however, popularized hypotheses propose that the physical constraints imposed by localized cell death inhibit pathogen

movement

and nutrient

are now

able

to cause

disease

in the previously

resistant

plant.

bacteria infect plants.

that bacterial gene products are ‘delivered

to’ and ‘perceived

pathways in

acquisition.

The plant’s recognition of a phytopathogenic bacterium is specified genetically by the presence of avirulence genes [3’] in the pathogen and complementary resistance genes in the host [4*,5]. All pathogens that have overcome host resistance have been shown to acquire mutations at their avirulence gene loci. Such mutations have resulted in the loss of avirulence function; however, because these bacteria have maintained their pathogenicity determinants, they

Phytopathogenic the /zq cluster

bacteria require to elicit the HR

the and

gene products of to be pathogenic.

Conserved amongst Gram-negative bacteria, /z@ clusters have been extensively studied in Erreinia amylovora (fire blight of apple and pear), Pseudomonas syringae pvs. syringae and tomato (leaf spot of bean, tomato and Arabidopsis), Ralstoniasolanaceanrm (bacterial wilt of tomato and potato), and Xanhomonas campestris pv. vesicatoria (bacterial spot of pepper and tomato) [6’]. These Rq clusters have been subdivided into two groups based on gene homology, organization and regulation: Erw’nia and Pseudomonas clusters comprise group I and Ralstonia and Xanthomonas clusters comprise group II [7**]. The pivotal observation that the proteins encoded by /r~ genes are homologous to those encoded by the virulence genes of several mammalian pathogens (Yersinia, Shigella and Salmonella) has revealed that Hrp proteins may comprise a type III protein secretion system capable of delivering proteins to plant cells [8]. Furthermore, the exciting discovery that /IQ genes and avirulence genes are co-regulated [9,10*,11*] suggests that avirulence determinants may also be secreted by the type III pathway. Recognition of these determinants is, thus, expected to occur inside plant cells by resistance gene products, either directly or indirectly

[4',51. Here we discuss briefly protein signaling in phytopathogenie bacteria and the role of the type III secretion pathway in pathogenesis and the elicitation of plant defense (Table 1; Figure 1).

Proteins secreted by the type III pathway Harpins were the first proteins found to be secreted in a &-dependent manner from cultures of E. amylovora [l&13], R. solanaceantm [14] and Z? syringae [15] (Table 1). This discovery confirmed the existence of a type III, /iqencoded, secretion pathway in phytopathogenic bacteria. The absence of any amino-terminal signal sequence in harpins is consistent with proteins shown to be secreted by the type III pathway in mammalian pathogens [8]. Indirect immunofluorescence microscopy has localized HrpZ from p syringae pv. syringae to the cell wall of tobacco cells [16] suggesting that cell wall association may be a requirement for harpin-induced signal transduction in plants. Mutational analyses have surprisingly revealed that most harpins do not directly control host specificity or elicitation of the HR [6’]. The role of harpins, thus, remains elusive. Recent studies leading to the discovery of a new class of pectate lyases (class III) are providing new insights.

110

Host-microbe

interactions: bacteria

Table 1 Proteins secreted

by the type III pathway

Protein classes

in phytopathogenic

bacteria.

Bacterium

Protein designation

Role in virulence

E. amylovora

Avirulence

E. amylovora E. carotovora E. chrysanthemi E. stewafl;; I? syringae pv. glycinea I? syringae pv. syringae P syringae pv. tomato

Harpinsf

Class III pectate

lyases

Pilins

DspE(A)’

Yes

HrPNE,

Yes

HrPNE,, HrPNEch

Yes

No

HrPNE,

No

HrpZp,,

Yes

H@p,,

No

HrpZpJEXP-45)

No No

/? solanacearum

PopA

E. amylovora

HrpW* HrpW

No

P. syringae pv. syringae f? syringae pv. tomato

HrpA (EXP-10)s

Yes

No

*DspE(A) is homologous to AvrE from P. syringae pv. tomato. fHarpins are glycine-rich, cysteine-lacking polypeptides. It should be noted that PopA also has avirulence-like properties. Comprehensive discussions reviewing harpins have recently been published [6’,7**]. fHrpW polypeptides possess an amino-terminal harpin-like domain and a carboxy-terminal pectate lyase domain. SHrpA shares no homology with other polypeptides; however, it appears to be a structural component of an hrp-encoded pilus.

The ir7pIY genes identified in P sytingae pv. tomato (A0 Charkowski, JR Alfano, G Preston, J Yuan, SY He, A Collmer, personal communication) and E. amylovora (JF Kim, CH Zumoff, SV Beer, personal communication) encode polypeptides with amino-terminal harpin-like domains and carboxy-terminal pectate lyase-like domains. Curiously, these polypeptides do not possess known pectate lyase enzymatic activities; however, both HrpW proteins have been shown to be secreted, suggesting that they may interact with pectic fractions in plant cell walls (A0 Charkowski, JR Alfano, G Preston, J Yuan, SY He, A Collmer, personal communication; JF Kim, CH Zumoff, SV Beer, personal communication). The striking similarities between class III pectate lyases and harpins raise many questions regarding their function(s) at the bacterial-plant cell wall interface and their evolution in phytopathogenic The hq-dependent from I! sytingae

bacteria. secretion pv. tomato

of five extracellular proteins has also been observed (des-

Work aimed at understanding the mechanism of HrpZ secretion has revealed that altered localization of this harpin in II syringae pv. sytingae Arp mutants is controlled by specific components in the type III pathway. Two /Iq operons, hpJ and /ilpr/, were shown to assist HrpZ translocation across the bacterial inner membrane, whereas the &C operon was necessary for its translocation across the outer membrane [‘ZO’]. Clearly, the dissection of &&mediated protein signaling is only just beginning. However, a working hypothesis is that plant-pathogenic bacteria utilize a protein complex in their envelopes, and possibly another complex at the bacterial-plant interface, to translocate elicitors directly to the plant cell (Figure 1).

Role of avirulence In general, topathogenic avirulence revealing campestris

genes?

the function of avirulence bacteria remains unclear.

proteins in phyThe majority of

genes encode novel polypeptides with no features. Sequence analysis of AvrBs2 from X. pv. vesicatoria, however, has provided clues to its

ignated EXP-10, EXP-22, EXP-43, EXP-45 and EXP-60 according to molecular weight) [17]. hpA gene encodes EXP-10, which forms a filamentous surface appendage (pilus) on solid surfaces under @-inducing conditions

biochemical function. AvrBs2 shares sequence similarity with agrocinopine synthase from Agrobacterium rumefaciens and UgpQ from Escheric/ria co/i [Zl]. Although these proteins have different functions, both appear to modify

[ 180.1. A nonpolar hpA mutant incapable of forming a pilus was unable to elicit the HR or cause disease in plants. This exciting discovery indicates that bacterial attachment to plant cells may be required for the transport of proteins across the plant cell wall. Interestingly, pilus assembly by Agrobacterium tumefaciens is required to transfer T-DNA and the VirE2 protein into tobacco cells [19]. EXP-45 and EXP-60 were shown to encode HrpZ [17] and HrpW (A0 Charkowski, JR Alfano, G Preston, J Yuan, SY He, A Collmer, personal communication) respectively. The identities of the other EXPs await further characterization.

phosphodiester

linkages.

The only avirulence protein with a proposed role in the synthesis of a bacterial elicitor is AvrD from P sytingae pv. tomato. E. co/i carrying avrD release syringolide compounds in culture that are active in causing a HR in soybean RPG4 cultivars [ZZ]. Biochemical studies have identified a syringolide-binding protein in soybean not specified by the RPG4 locus [23]. Thus, recognition of the avrD-specified elicitor appears to occur prior to RPGCmediated signal transduction in soybean.

Protein

signaling

via type

III secretion

pathways

in phytopathogenic

bacteria

Mudgett

111

and Staskawicz

Figure 1

dellvery -ectly

(b)

Bacterial

evidence

w

Plant evidence

Plant cell fig*

/

/

1 *vrW2

1-l-R

\\

Klnase

>

HR

casade

Current Op,n,on I” M,crob,ology

Models depicting the current evidence for the delivery of bacterial proteins to plant ceils via the type Ill secretion pathway in phytopathogenic bacteria. (a) The scanning electron micrograph shows /? syringae pv. tomato multiplying in the intercellular spaces of leaf mesophyll tissue of Arabidopsis. (b) Bacterial evidence summarizing the many genetic and biochemical analyses investigating the functional roles of Hrp proteins in the secretion of harpins, class Ill pectate lyases (HrpW), avirulence determinants (DspE[Al), and unknown proteins (EXP-22 and EXP-43). The Hrp proteins are designated as C, J, R, S, T, U and pilus for simplicity (see references [6’,7”1 for comprehensive discussions). PrhA is the first bacterial protein shown to be regulated in an hrp-independent, plant-dependent manner. (c) Plant evidence confirming that the molecular recognition of avirulence proteins occurs inside plant cells. Bacterial avirulence cxcles, respectively. RPSP, RPM1 and PTO are plant resistance proteins.

proteins

and plant proteins

are represented

by squares

and

112

Host-microbe

interactions: bacteria

For years, plant pathologists have tried to use biochemical approaches to identify enzymatic functions for avirulence proteins. Numerous groups have isolated cell-free extracts from induced bacteria possessing avirulence to characterize an active elicitor fraction.

genes, hoping Unfortunately,

none of the extracts triggered a HR when they were infiltrated into the plant apoplast. It was speculated that avirulence elicitors were unstable and present at low levels evading detection. However, purified avirulence protein preparations similarly failed to elicit HR phenotypes. The

demonstration

that /Ilp genes

and avirulence

genes

are co-regulated in nonpathogenic bacteria finally linked the type III secretion pathway and avirulence proteins with the elicitation of the HR. E. co/i carrying the li syringae pv. syringae hp gene cluster was able to phenotypically express avrA, avrPto, avrRpm1, avrRpt2, and avrPph3 in plants [lo’]. Similar results were observed with P~%~orescens carrying the same /17p cluster and a-vrB [ll’]. Importantly, this work disclosed that avirulence elicitors may be directly delivered inside plant cells via the ir7p-encoded type III secretion apparatus. Thus, avirulence proteins presented to the outside of plants presumably failed to elicit the HR because they were incapable of penetrating the plant cell wall. The secretion of a newly identified avirulence protein, DspE(A), from E. amylovora (see below) supports the secretion of avirulence proteins and this model. It is interesting to note that AvrRxv from X. campesttis pv. vesicatoria shares homology with three secreted proteins, AvrA from S. enterica, YopJ from I: pseudotuberculosis and YopP from I: enterocolitica [24,25]. If avirulence

elicitors

are targeted

to plants

revealing

their

presence, why would bacteria retain these genes? Most avirulence genes have no apparent role in pathogenicity; however, five genes (avrBs2, au-66 and pthA from X. campestris, and avrE and avrRpm/ from i? syringae) have displayed positive virulence functions when gene knockout mutants have been assayed for virulence [3’]. This suggests that pathogens actually use these proteins during the infection process for bacterial fitness or even virulence. One hypothesis is that avirulence proteins may contribute to growth switch from epiphytic

adaptation in plants when bacteria growth to intercellular growth [26].

death. Agrobacrerium-mediated transient gene expression was concurrently used to demonstrate the recognition of AvrPto from P sytingae pv. tomato in resistant Pto tobacco plants [28’,29*], and the recognition of AvrBs3 from X. campestris pv. campestris in resistant

Bs3 pepper

plants

[30*].

The yeast two-hybrid system was then employed to show a direct physical interaction of an avirulence protein with a resistance protein, namely AvrPto with Pto [28*,29*]. Although yet been

an interaction demonstrated

between these proteins has not in vivo, mutations that have

disrupted AvrPto-Pto interactions in yeast have also abolished disease resistance in plants [28*,29*]. AvrPto is now modeled to interact with Pto in the plant cytoplasm resulting in the activation of a kinase cascade involving Ptil, Prf, Pti4, PtiS and Pti6, to induce downstream defense genes [31,32]. Although the subcellular localization of AvrRptZ, AvrRpml and AvrPto remains to be determined, members of the Xanthomonas avirulence/pathogenicity family appear to be transported to the plant nucleus. Functional nuclear localization signals (NLS) have been identified at the carboxy-terminal regions of Avrbb, PthA and AvrBs3 [30*,33]. Nuclear targeting was demonstrated by fusing putative signal sequences with the B-glucuronidase gene and transiently expressing the reporter constructs in onion epidermal cells using biolistic bombardment [30’,33]. Mutational analyses showed that AvrBs3 lacking all NSL remains cytoplasmic in onion cells and fails to elicit cell death in pepper [30’]. Thus, recognition and translocation of AvrBs3 to the host nucleus appears to be required for induction of this HR. It is intriguing that a recent screen of a yeast two-hybrid library made from pepper cDNA has identified a specific AvrBs3-binding protein highly homologous to importin-a (B Szurek, G Van den Ackerveken, U Bonas, personal communication). This interaction was specific for the peptide region of AvrBs3 containing the NLS. We await the cloning of the pepper resistance gene Bs3 for further elucidation of the role of AvrBs3 in signaling cascades in planta. Because

transient

expression

of avirulence

proteins

in

Evidence connecting avirulence function with the type III secretion pathway opened the door to investigations verifying that the molecular recognition of avirulence

resistant plants leads to cell death, biochemical and cell biological dissection of this response has been difficult. Researchers are now developing powerful genetic tools to study the conditional expression of avirulence genes in planta. One successful approach has led to the construction of transgenic Arabidopsis plants containing avrRpt2 gene

proteins occurs inside plant cells. Two transient expression assays were developed to show that expression of avirulence polypeptides in planta was sufficient to elicit a resistance gene-dependent HR. Biolistic bombardment was used to introduce the DNA of two I? syringae avirulence genes, avrB and avrRpt2, into Arabidopsis leaves containing the complementary resistance genes, RPM/ and RPSZ, respectively [ 11’,27*]. Both AvrB and AvrRpt2 were biologically active inside plant cells resulting in cell

under the control of a tightly regulated, glucocorticoidinducible promoter (TW McNellis, MB hludgett, I( Li, T Aoyama, D Horvath, N-H Chua, BJ Staskawicz, unpublished data). Dexamethasone-induced expression of avrRpt2 in the transgenic lines having the RPST resistance gene caused a cell death response resembling a P s_)&gae-induced HR. A high level of avrRpt2 expression in susceptible transgenic plants ultimately resulted in cell death revealing a virulence role for this protein (TW

Role of avirulence

proteins in planta

Protein

signaling

via type III secretion

McNellis, MB Mudgett, K Li, T Aoyama, D Horvath, N-H Chua, BJ Staskawicz, unpublished data). These transgenic plant lines have been mutagenized and are currently being screened for defects in the avr&tZ-specified cell death pathway. The characterization of isolated mutants is expected to yield new host factors required for both resistance

and virulence

responses.

New virulence determinants Transposon uncovered

mutagenesis new virulence

in phytopathogenic

function

of in to

encode a polypeptide displaying significant similarities to TonB-dependent outer membrane siderophore receptors [34**]. Mutations in prhA abolished activation of /rrpB and additional /zrp genes when bacteria were co-cultured with Ambidopsis cells, whereas the expression of these genes in lIq-inducing minimal media was not affected. These results suggest that PrhA is a receptor for plant-derived signals. Significantly, this work provides the first example of a specific plant-dependent pathway controlling the induction of /Iq genes. In E. amylovora, transposon mutagenesis has identified a disease specific (dsp) locus that is not required for the HR but is essential for fire blight symptoms [3.5,36*]. A two-gene operon responsible for this phenotype was simultaneously characterized, and was designated dspEF [35] and dspAB [36’]. These genes are homologous with genes in the avrE locus of I? sytingae pv. tomato [37]. DspE(DspA) and AvrE are large, hydrophilic proteins, whereas DspF(DspB) and AvrF are small, acidic proteins resembling chaperones for virulence factors secreted by type III pathways in mammalian pathogens [38]. Interestingly, DspE was found secreted into the media of induced E. amylovora culture [39”] (Table 1). Subsequent studies showed that p syringae pv. glycinea carrying the dspEF operon elicited a HR on soybean, demonstrating that this pathogenicity operon can also function as an avirulence locus [35]. This work nicely describes the first pathogenicity locus in E. amylovora encoding a secreted polypeptide possessing both virulence and avirulence properties. It should also be noted that monogenic resistance to fireblight has not yet been reported. Recognition of DspE in soybean indicates the existence of a resistance gene. Cloning this resistance gene may lead to protection against fire blight in the field.

Conclusions One of our next challenges is the acquisition of direct evidence for the hip-dependent delivery of bacterial proteins to plant cells. If phytopathogenic bacteria use the type III secretion pathway to infect plants, then bacterial protein translocation and localization in planta must be verified. Furthermore, potential intracellular host targets need to be identified. Plant resistance proteins are expected to

bacteria

as receptors

for

Mudgett

bacterial

113

and Staskawicz

avirulence

elicitors;

however, a direct interaction has been shown in only one case. Several attempts to demonstrate interactions between avirulence proteins and their complementary resistance proteins have failed. Researchers will need to rely on new cell biological and biochemical approaches to directly test these models. Such studies will probably be best accomplished using p syringae and Arabidopsis, two genetically

of phytopathogenic bacteria has determinants unlinked to the /jlp

cluster. In R. solanacearum, thepr/rA genre (plant regulator hrp genes) was found to control interactions with plants a HrpB-independent manner [34**]. pr/lA is predicted

pathways

tractable

systems.

If bacterial proteins are delivered to plant cells, then what are their roles in phytopathogenesis? How many are delivered to the host? Which proteins proteins specify host resistance versus nonhost resistance? Are chaperones required? What are the signals that specify protein secretion? Are the secretion determinants in proteins from phytopathogenic bacteria encoded in their messenger RNA, as was determined for Yop proteins from Z: enrerocolitica [40”]? How universal are the mechanisms for type III secretion pathways? Can we assume that plant and animal pathogens use similar mechanisms to infect different hosts? Answers to these questions will strengthen our understanding of the bacterial type III secretion machineries and the specific mechanisms controlling phytopathogenesis.

Acknowledgements ‘l‘hanks to I\lanucl thanks Collmcr the

Sainz

CO hlarie-Anne for sharing

National

for

manuscripts

Insrirute

readmg

uf

the

prior

to publication.

manuscript.

hlBh1

is supoortcd

NRSA grant 1 F32 Gh118414-01. of Energy grant DE-FG03-88ER13917.

and recommended

Special

Boucher and Alan

of Health

suppnrtcd by chc IIcpartment

References

critical

Barn): Steve Beer, Christian

by

BJS is

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. ..

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Lee CA: Type III secretion systems: machines to deliver bacterial proteins into eukaryotic cells? Trends Microbial 1997, 5:148-l 56.

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23.

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Kearney B. Roy M, Staskawicz BJ: mutations in a single open reading and avirulence in Xanthomonas avr6s2. J Bacterial 1996, 178:4661-

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13.

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28. .

14.

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15.

He SY, Huang H-C, Collmer A: Pseudomonas syringae pv. syringae harpinpss: a protein that is secreted via the hrp pathway and elicits the hypersensitive response in plants. Cell 1993, 73:1255-l 266.

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19.

20. .

Scofield SR, Tobias CM, Rathjen JP, Chang JH, Lavelle DT, Michelmore RW, Staskawicz BJ: Molecular basis of gene-forgene specificity in bacterial speck disease of tomato. Science 1996, 274:2063-2065. See annotation [29’1. 29. .

Tang X, Frederick RD, Zhou J, Halterman DA, Jia Y, Martin GB: Initiation of plant disease resistance by physical interaction of AvrPto and Pto kinase. Science 1996, 274:2060-2063. This work, along with [28*] provides the first data supporting a physical interaction between a bacterial avirulence protein and a plant disease resistance protein. 30. .

Van den Ackerveken G, Marois E, Bonas U: Recognition of the bacterial avirulence protein AvrBs3 occurs inside the host plant cell. Cell 1996, 87:1307-l 316. This work, in addition to [l 1’,27’-29’1, confirms that molecular recognition of an avirulence protein occurs inside a plant cell. 31.

Zhou J, Tang X, Martin GB: The pto kinase conferring resistance to tomato bacterial speck disease interacts with proteins that bind a &-element of pathogenesis-related genes. EM60 J 1997, 16:3207-3218.

32.

Salmeron JM, Oldroyd GED, Rommens CMT, Scofield SR, Kim H-S, Lavelle DT, Dahlbeck D, Staskawicz BJ: Tomato Ptf is a member of the leucine-rich repeat class of plant disease resistance genes and lies embedded within the Pto kinase gene cluster. Cell 1996, 66:123-l 33.

33.

Yang Y, Gabriel DW: Xanthomonas avirulence/pathogenicity gene family encodes functional plant nuclear targeting signals. MO/ P/ant-Microbe lnreract 1995, 8:627-631,

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Charkowski AO, Huang H-C, Collmer A: Altered localization of hrpZ in Pseudomonas syringae pv. syringae hrp mutants suggests that different components of the type Ill secretion pathway control protein translocation across the inner and outer membranes of Gram-negative bacteria. J Bacterial 1997, 179:3866-3874. Detailed mutational and biochemical analyses of the hrp gene cluster providing insight into the functional roles of Hrp proteins in protein secretion.

34. ..

Marenda M, Brito B, Callard D, Genin S, Barberis P, Boucher C, Arlat M: PrhA controls a novel regulatory pathway required for the specific induction of Ralstonia solanacearum hrp genes in the presence of plant cells. MO/ Microobiol1998, in press. Significantly, this work provides the first example of a specific plant-dependent pathway controlling the induction of bacterial hrp genes. 35.

Bogdanove AJ, Kim JF, Wei Z, Kolchinsky P, Charkowskl AO, Conlin AK, Collmer A, Beer SV: Homology and functional

Protein

signaling

via type III secretion

similarity of a hrp-linked pathogenicity operon, dspEF, of Erwinia amylovora and the avrE locus of Pseudomonas syringae pathovar tomato. Proc Nat/ Acad SC; USA 1998, in press. Gaudrlault S, Malandrin L, Paulin J-P, Bamy M-A: DspA, an . essential pathogenicity factor of Erwinia amy/ovora showing homology with AvrE of Pseudomonas syringae, is secreted via the Hrp secretion pathway in a DspB-dependent way. MO/ Microbial 1997, 26:1057-l 069. This work describes the identlflcation of a new bacterial determinant that is secreted by the type III pathway that plays a role in phytopathogenicity. 36.

37.

Lorang JM, Keen NT: Characterization of avrE from Pseudomonas syringae pv. tomato: a hrp-linked avirulence locus consisting of at least two transcriptional units. MO/ PlantMicrobe interact 1995, 8:49-57.

pathways

38.

in phytopathogenic

bacteria

Mudgett

and Staskawicz

115

Comelis GR, Wolf-Watz H: The Yersinia Yop virulon: a bacterial system for subverting eukaryotic cells. MO/ Microbial 1997, 23:861-867.

39. ..

Bogdanove Al, Bauer DW, Beer SV: Erwinia amylovora secretes DspE, a pathogenicity factor and functional AvrE homolog, through the hrp (type Ill secretion) pathway. I Bacferiol 1989, in press. These researchers nicely describe the first pathogenicity locus encoding a secreted polypeptide possessing both virulence and avirulence properties. 40. ..

Anderson DM, Schneewind 0: A mRNA signal for the type Ill secretion of Yop proteins by Yersinia enterocolitica. Science 1997, 278:l 140-l 143. A landmark discovery revealing that the signal targeting bacterial proteins through the type Ill secretion pathway is not encoded by amino acid sequence or protein structure. Rather, the signal is specified by the mRNA structure just upstream of the start codon for secreted Yop proteins.