Mikael Göransson

Upstream activating sequences that are shared by two divergently transcribed operons mediate cAMP-CRP regulation of pilus-adhesin in Escherichia coli

Transcription of the genes encoding pilus‐adhesin of serotype F13 in digalactoside‐binding Escherichia coli required activation by the cAMP‐CRP complex. Analysis of protein‐DNA interaction in vitro showed that CRP bound in a cAMP‐dependent manner to a sequence located 0.2 kb upstream of the point of transcription initiation of the pilus subunit operon. The cAMP‐CRP activation included, in addition to the main pilus operon, the oppositely oriented operon encoding the Papl regulatory protein. Furthermore, the auto‐regulatory product of the promoter‐proximal gene (papB) in the pilus subunit operon was found to stimulate the papl transcriptional unit. Thus the cAMP‐CRP complex and PapB might act in concert and indirectly promote pili synthesis by stimulating expression of the Papl positive regulator. The results of trans complementation experiments and analyses using lacZ operon fusion derivatives showed that the cAMP‐CRP activation also operated directly in cis on the pilus subunit operon. The region containing the CRP binding site appeared to function as an upstream activating sequence since deletion abolished expression even when the pap regulatory proteins Papl and PapB were supplied in trans. The implications for possible mechanisms of transcriptional activation by the cAMP‐CRP complex at this novel location between the two oppositely oriented operons are discussed.

Regulatory Interactions among adhesin gene systems of uropathogenic Escherichia coli

ABSTRACT Uropathogenic Escherichia coli strain J96 carries multiple determinants for fimbrial adhesins. The regulatory protein PapB of P fimbriae has previously been implicated in potential coregulatory events. The focB gene of the F1C fimbria determinant is highly homologous to papB; the translated sequences share 81% identity. In this study we investigated the role of PapB and FocB in regulation of the F1C fimbriae. By using gel mobility shift assays, we showed that FocB binds to sequences in both the pap and foc operons in a somewhat different manner than PapB. The results of both in vitro cross-linking and in vivo oligomerization tests indicated that FocB could function in an oligomeric fashion. Furthermore, our results suggest that PapB and FocB can form heterodimers and that these complexes can repress expression of the foc operon. The effect of FocB on expression of type 1 fimbriae was also tested. Taken together, the results that we present expand our knowledge about a regulatory network for different adhesin gene systems in uropathogenic E. coli and suggest a hierarchy for expression of the fimbrial adhesins.

Genes Determining Adhesin Formation in Uropathogenic Escherichia coli

The ability to adhere to the site of infection is considered to be a prerequisite for most bacterial infections. For a number of gram-negative species there is evidence that filamentous surface structures on the bacteria, called pili or fimbriae, are the adhesins that bind to epithelial cells of the host. The adhesins interact with complementary receptors which are carbohydrate-containing polymers on the target cells. The importance of adhesion in pathogenesis of infectious disease has stimulated intensive research aimed at elucidating the molecular basis of such interactions. It is hoped that an increased knowledge about bacterial adhesion will facilitate more rational developments of therapeutic or preventive measures against a given pathogen. Analysis of genetic determinants has provided a great deal of insight into the biogenesis of adhesins. By molecular cloning techniques, in vitro mutagenesis, DNA sequencing, and other procedures, individual gene products are identified and their functions assessed. Studies of regulatory features help our understanding of what molecular mechanisms determine expression of the virulence function.

Functional and structural homology among regulatory cistrons of pili-adhesin determinants in Escherichia coli

SummaryExpression of the digalactoside-binding Pap pili involves two trans-acting regulatory genes, papB and papI. Using pap-lac operon fusions and DNA hybridization probes derived from pap DNA we tested whether or not other pili-adhesin determinants from different Escherichia coli strains encode homologs to the pap regulatory genes. Digalactoside-specific clones of serotypes F72 and F11 complemented papB and papI mutants of the Pap (serotype F13) clone and DNA hybridization analysis showed that the clones are homologous in the DNA sequences encoding the two regulatory genes. Similar results were obtained with an S-pili determinant which mediates binding to sialic acid-containing receptors and the findings suggest that the regulatory regions may be more conserved than other genes in different pili-adhesin gene clusters. Determinants for type 1-pili (mannose-specific binding) and for pili associated with enterotoxigenic E. coli (K88, K99, CFAI, CFAII) did not appear to contain DNA sequences homologous to papB or papI. E. coli strain J96, which was the origin of the pap DNA, was found to carry two additional copies of papB-papI homologous sequences in the chromosome. In strains expressing more than one kind of pili the trans-active gene products thereby may allow for regulatory interaction between separate pili-adhesin gene systems.

Genetics of Histone-Like Protein H-NS/H1 and Regulation of Virulence Determinants in Enterobacteria

Isolates of E. coli associated with intestinal or with extraintestinal disease are often characterized by their ability to express different properties thought to contribute to bacterial virulence. Examples of such properties are the synthesis of various types of adhesins, production of different kinds of cytotoxins (e.g. enterotoxins and hemolysins), ability to invade host tissue cells, and expression of certain capsule- and O-antigens. Changes in growth conditions may profoundly affect how different virulence-associated properties are expressed. The type of growth substrate, osmolarity, and growth temperature are examples of environmental factors found to influence the phenotypic expression of some of the virulence associated properties monitored under laboratory conditions. We have studied the expression of E. coli adhesins typically produced by many isolates from urinary tract infections in man. The P-specific adhesins are fimbrial adhesins, also referred to as pili, and they mediate binding to α-D-Gal-(1–4)-β-D-Gal-containing glycolipid structures. Genetic studies of the pap gene cluster have revealed functions of several of the gene products required for biogenesis of this type of adhesin (see Tennent et al., 1990 for a recent review).

Biogenesis of Galα(1–4)Gal binding P-pili of Uropathogenic E. Coli

Escherichia coli is the most common cause of urinary tract infections (UTI). Depending on the virulence of the organism and the efficiency of the host defense system, these UTI appear either as asymptomatic bacteriuria, acute cystitis or acute pyelonephritis. A number of putative virulence properties have been recognized in E. coli isolates associated with acute pyelonephritis in otherwise uncompromised children. One such property is the ability to express mannose-resistant (MR) hemagglutinins. Most E. coli are capable of forming type 1 pili that carry a mannosespecific adhesin. Pyelonephritogenic E. coli additionally carry one or more chromosomal gene clusters that encode pili associated with MR hemagglutination. Of the several MR hemagglutinins that are known today, the best characterized is the Galα(1–4)Gal-specific adhesin associated with P-pili. The name P-pili refers to the observation that this adhesin acts as a hemagglutinin for erythrocytes which express any of the P blood group antigens.