Assembly and adhesive properties of curli : A stationary phase-specific surface organelle in gram-negative enteric bacteria

Sammanfattning: Assembly and adhesive properties of curli: A stationaryphase-specific surface organelle in gram-negative enteric bacteria. Mårten Hammar The natural environments of bacteria are most often characterizedby suboptimal conditions for growth, e. g., limited supply ofnutrients. Consequently, periods of dormancy or negligible growth arethe rule rather than the exception. Starvation leads to profounddevelopmental changes. The most dramatic response to starvation isthe generation of dormant spores by many species of gram-positivebacteria and the generation of multicellular fruiting bodies bymyxobacteria. In contrast, many gram negative bacteria, e. g.,enteric bacteria like Escherichia coli and Salmonellaenterica, do not enter a dormant state but maintain a low levelof metabolism combined with an increased resistance to environmentalstresses. Although not considered a typical starvation-inducedresponse, bacteria express a variety of surface structures inresponse to diverse environmental conditions. Often, these structuresare proteinaceous filaments extending out from the bacterial cellsurface. These structures serve the purpose of mediating contactbetween the bacterium and a eukaryotic cell surface, a tissue matrixor serum protein, or to other bacteria, conspecific or of otherspecies. These interactions are often the commited steps leading tosubsequent colonization of an epithelial surface, entry into a hostcell, exchange of DNA between bacteria, or development of a bacterialcommunity organized as biofilms, colonies or multicellular fruitingbodies. In Escherichia coli, a novel type of fimbriae-likestructure denoted curli was recently discovered. This surfaceorganelle is characterized by a distinct morphology, a high bindingaffinity for many eukaryotic proteins, and a unique pattern ofstationary phase-dependent expression. To further understand the mechanisms of curli biogenesis and theregulation of curli expression, a genetic analysis was performed toidentify the genetic determinants involved in these processes. Thisthesis presents the identification of six genes, csgDEFG and csgBA,the products of which are specifically involved in curli production. CsgA is the major subunit protein of the curli filament. CsgB is asurface-exposed protein required for polymerization of CsgA subunitsinto curli filaments and for anchoring of these to the cell surface.CsgG is an outer membrane-anchored periplasmic lipoprotein whichstabilizes the CsgB and CsgA proteins. CsgF, together with CsgB,participates in nucleation of CsgA subunits. A cell deficient innucleation function secretes soluble CsgA monomers or oligomers tothe growth medium. These CsgA subunits are polymerization-competent,i. e., they spontaneously polymerize to curli fibers on the surfaceof any cell presenting a functional nucleator (CsgB). This process ofextracellular formation of curli indicates a novel pathway offimbriae biogenesis, the extracellular nucleation-precipitationpathway. The addition of subunits to the growing filament seems to bedriven by mass action and guided only by the diffusion gradientbetween the source of secreted subunits and the growing curli tip. The adhesive properties of curli are dependent on CsgA subunitspolymerized in the presence of CsgE. Thus, inactivation of the csgEgene results in curli deficient in binding. A similar type ofadhesion-deficient curli is produced by cells devoid of Nacetylglucosamine-6-phosphate deacetylase. Several regulatory proteins controlling the expression of curliwere identified. CsgD is required for transcription of the csgBAoperon. OmpR, a transcriptional activator responding to osmolarity,and d, a stationary phase-specific sigma factor, activatestranscription of the csgDEFG operon. Mutations in several other lociaffect transcription of the csg genes, indicating a complexregulation of curli expression where different signals are mtegrated. Key words: Escherichia colil/curli/ fimbriae/ csg/starvation ISBN 91-628-2605-0