Identification and characterization of genes determining receptor binding and pilus length of Escherichia coli type 1 pili

Abstract

We describe the identification and characterization of two genes and their gene products responsible for determining receptor binding and pilus length in type 1-piliated Escherichia coli. One gene, pilE, conferred the ability of piliated cells to agglutinate guinea pig erythrocytes. The other gene, pilF, determined pilus length, in that mutants having lesions in pilF had very long pili. The two genes were detected after TnS mutagenesis of a cloned segment of DNA that normally complemented a pilE lesion in the chromosome. Thus, lesions in pilE or pilF on the cloned segment resulted in mutants having the PilE-phenotype (piliated but unable to agglutinate erythrocytes). Introduction of the plasmid-encoded mutant alleles of pilE and pilF into the chromosome followed by electron microscopic examination of the mutants showed that only lesions in pilF conferred the striking increase in pilus length. Mutations in pilF could be complemented in trans by the original cloned segment to produce cells with parental-length pili. Minicell transcription and translation of the cloned pilE and pilF genes having representative TnS insertion mutations showed that the pilE gene product was a protein of ca. 31 kilodaltons and that the pilF gene product was a protein of ca. 18 kilodaltons. We believe that the pilF gene product may act as a competititve inhibitor of pilus polymerization. Thus, pilus length may be controlled by the ratio of pilin to pilF gene product present within the cell.Type 1 pili of Escherichia coli provide a convenient model for studying the control and assembly of supramolecular extracellular structures. Also, they provide a model for cell-cell interaction, since type 1 pili are required to mediate attachment of E. coli to a variety of eucaryotic cells through a mannose-sensitive interaction with a receptor on the eucaryotic cell (4).One interesting facet of the assembly of type 1 pili concerns how the cell monitors the number of pili it has and how long those pili are. These two considerations would seem to be important for the cell to conserve energy. However, since pili are on the outside of the cell, it is unclear how the cell would keep track of these two parameters. It is apparent from recent work that at least one mechanism exists for controlling the number of pili per cell. This fact is indicated by the existence of hyperpiliated mutants (13). Since hyperpiliated mutants also have longer pili (13), one might infer that the cell has a way to monitor pilus length also. However, no such pilus length mutants have been isolated.Another interesting feature of type 1 pili concerns their involvement in receptor-ligand interactions. Recently, it has become evident that while type 1 pili are required for receptor binding, their presence is not sufficient for binding (8). This fact is indicated by the existence of fully piliated mutants that fail to agglutinate guinea pig erythrocytes (8).

Immunochemical and functional studies of Actinomyces viscosus T14V type 1 fimbriae with monoclonal and polyclonal antibodies directed against the fimbrial subunit

“…analogous to the various adhesins a.,soci;tted L+I'!? c!lfTerent E. coli pili (Lindberg et al, 1986;Moch et d., 1987;Maurer & Orndorff, 1987;Abraham et af., 1987). Further studies of A .…”

Mast cell degranulation induced by type 1 fimbriated Escherichia coli in mice.

“…Plasmid pJP4 encodes the E. coli type 1 fimbrial chaperone, FimC (15,24). The properties of each of these plasmids have been described in detail elsewhere (15,17,(21)(22)(23)(24) (26). The ( 14,15).…”

“…We have shown that type 1 fimbriae promote bacterial binding and activation of different immunoinflammatory cells such as lymphocytes and neutrophils, evoking a variety of biological reactions in the activated cells ( 14,15). The determinant of adherence and host cell activation on type 1 fimbriae is a minor protein, FimH, which is located in a functionally competent configuration at the tips of each fimbrial filament (16)(17)(18)(19).…”

“…The determinant on type 1 fimbriae responsible for mast cell degranulation and histamine release was FimH, the mannose-binding subunit. Bacterial strains and culture conditions E. coli ORN103(thr-l leu-6 thi-lA(argF-lac) U169xyl-7 ara-13 mti-2 gal-6 rpsL tonA2fhuA2 minA minB recA13 6(fimABCDEFGH) (17,21) (23). Plasmid pRT4 was subcloned from pSH2 and contains only thefimH gene (15).…”

FimH family of type 1 fimbrial adhesins: functional heterogeneity due to minor sequence variations among fimH genes

“…Approximately 1,000 copies of the 17-kDa primary structural subunit, FimA (or PilA), are polymerized into a right-handed helical fibril also containing minor amounts of the FimF, FimG, and FimH subunits (20,24,27,32). The 28-kDa FimH subunit has been shown by direct and indirect tests to be the fimbrial lectin (1-3, 20, 21, 27, 29, 32, 36, 55).…”