Functions of F Pili in Mating-Pair Formation and Male Bacteriophage Infection Studied by Blending Spectra and Reappearance Kinetics

Novotny, Charles P.; Raizen, Eileen; Knight, William S.; Brinton, Charles C.

doi:10.1128/jb.98.3.1307-1319.1969

Cited by 25 publications

(14 citation statements)

References 21 publications

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“…(The rate of specific pair formation is defined as the percentage increase in number of specific pairs per unit time in relation to the maximum number of specific pairs attained.) We believe, therefore, along with other investigators (2,18,28,29), that the ability to form specific pairs is solely dependent on the presence of F pili and, furthermore, that it is little affected by differences in F pili length and other cellular structures or by capabilities resulting from the various cultural conditions employed.…”

Section: Relationship Between Growth Conditions Of Donorsupporting

confidence: 62%

“…These three events must precede transfer of the donor chromosome during conjugation. After the discoveries that donor cells possess a donor-specific antigen (30) and are able to adsorb donor-specific ribonucleic acid (RNA) phages (24,25) to filamentous external structures (7) called F pili (4), it was demonstrated that these F pili are synthesized under the control of the fertility factor F (2)(3)(4) constitute the F-specific antigen (20), and are obligatory for specific pair formation (2,18,28,29). Brinton's group (2-4, 28, 29) and others (18,21,32) have further suggested that F pili may function as conjugation tubes in addition to their role in specific pair formation.…”

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confidence: 99%

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Early Stages of Conjugation in Escherichia coli

et al. 1969

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We initiated these studies to learn more about the initial events during bacterial conjugation and to optimize conditions for their occurrence. We found that cells in donor cultures grown anaerobically prior to mating have (i) a higher mean number of F pili per cell, (ii) longer F pili, (iii) a higher probability of forming specific pairs with Fcells, and (iv) a faster rate of initiation of chromosome transfer than cells grown aerobically. The growth medium for the donor culture also influences these same parameters: a rich medium is superior to a completely synthetic medium. Starvation of donor cells in buffered saline or for a required amino acid results in (i) a loss of F pili, (ii) a loss in the ability of donor-specific phages to adsorb, (iii) a loss of ability to form specific pairs with Fcells and to yield recombinants, and (iv) an increase in recipient ability. These changes occur as a function of starvation time, and at rates which are dependent on the conditions of prior growth and starvation of the donor culture. Either treatment provides a rapid method for the production of Fphenocopies from donor cultures. Resynthesis of F pili by cells within a starved donor culture commences very soon after restoration of normal growth conditions, but full restoration of donor ability, as measured by recombinant yield, occurs at a slower rate. We found, along with other investigators, that F pili are essential for specific pair formation. We also found, however, that the presence of F pili is not sufficient for display of donor ability, nor is the absence of F pili enough for cells to exhibit recipient ability. This suggests, therefore, that one or more components, in addition to F pili, are necessary for the conversion of specific pairs to effective pairs (or for chromosome mobilization, or both) and for preventing donor cells from acting as recipients. On the basis of our results, we suggest optimal conditions for achieving high mating efficiencies.

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Section: Relationship Between Growth Conditions Of Donorsupporting

confidence: 62%

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confidence: 99%

Early Stages of Conjugation in Escherichia coli

et al. 1969

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“…Heated cells should also be useful for studying the reappearance of pili. Usually, blended cells that only produce about 60% of the control level of pili (3,12) are used to study reappearance (3,4,15,18,19). It might be better to use cells heated to 500C for this purpose, because they produce normal numbers of pili when the temperature is lowered to 37°C.…”

Section: Discussionmentioning

confidence: 99%

“…MATERIALS AND METHODS Bacteria and bacteriophages. F pili were studied on a strain of E. coli K-12 (ERW) that is F+ His+/His- (15). R17, a spherical RNA phage, and M13, a filamentous DNA phage, were used to study donor-specific phage adsorption, and R17 was used to label pili for electron microscopy.…”

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Effects of high temperature on Escherichia coli F pili

Novotny¹,

Fives-Taylor²

1978

J Bacteriol

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The effects of high temperatures (46 to 5000) on the production of F pili by Escherichia coli were studied by electron microscopy. Attached F pili rapidly disappeared at 48 and 500C but not at 460C. Free pili were not denatured at these temperatures. The pili that disappeared from the cells at 500C did not appear as free pili in the culture supematant fluid, indicating that the pili had retracted to the cell surface or into the cell. The adsorption of either R17 phage or F pili antibody to the sides of pili prevented retraction. The disappearance of pili was accompanied by a loss in the ability to adsorb R17 phage but not M13 phage, suggesting that the tip of a pilus remains exposed after retraction.F pili rapidly disappear from the surface of donor strains of Escherichia coli K-12 under certain conditions. These conditions include infection with filamentous, donor-specific phages (9,11); incubation in the presence of sodium cyanide (13) and arsenate (17); and temperature shifts from 37 and 250C (14). Marvin and Hohn (11) were the first to suggest that F pili could retract. They observed that F pili disappeared during Ff phage infection and suggested that the pili retract into the cell with the phage on their tips. Other types of pili may also retract. The production of I pili by E. coli cells is stimulated by I pili antibody, and it was suggested that antibody might cause an increase in attached pill by preventing retraction (10). The shortening of Pseudomonas aeruginbsa pffi after adsorption of RNA phage to the sides suggests that they retract to the point of attachment of the phage (2). Novotny and Fives-Taylor (13) suggested that the disappearance of F pili in the presence of cyanide and at certain temperatures was due to retraction and presented a dynamic model for the growth of pili that supposes that F pili elongate and retract continuously and that the number of F pili per cell and the average length of pili reflect the state of equilibrium between these two processes.Studies of the outgrowth of F pili indicate that outgrowth requires the synthesis of ATP (16, 17) and the synthesis of RNA (8). The synthesis of DNA (16) or the synthesis of protein (3, 16) is not required. Although the synthesis of DNA is not needed for the reappearance of pili, there is evidence that DNA may be involved (7,8).Very little is known about the requirements for retraction. Retraction does not require the synthesis of ATP because pili retract in the presence of energy poisons (13, 17), and it does not require the synthesis of RNA and protein because rifampin, streptolydigin, and chloramphenicol have no immediate effect on retraction in the presence of cyanide (8). Retraction is inhibited by chloramphenicol after 15 min, suggesting that retraction requires a protein whose pool size is limited (8).Retraction and outgrowth are both sensitive to temperature. The sudden decrease in the number of attached F pili that occurs when cells are cooled from 37 to 250C (14) suggests that outgrowth, but not retraction, is inhibited at 250C. ...

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“…The ability of a bacterial cell to act as a genetic donor is often correlated with the presence of sex pili on the cell surface (16). Most sex factors specify sex pili which belong to one of two main classes, F-like or I-like (14), but other classes have recently been observed (6).…”

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Purification of Sex Pili from Escherichia coli Carrying a Derepressed F-Like R Factor

Beard¹,

Howe²,

Richmond³

1972

J Bacteriol

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A procedure for the purification of sex pili is described. Escherichia coli K-12 carrying Rldrdl9 was grown in nutrient broth and blended at the time of peak sex pilus production. The cells were removed by centrifugation, and the supernatant fraction was concentrated, dialyzed, and clarified in an ultrafiltration system. After an additional blend and a clearing spin, the material was centrifuged in a CsCl gradient, and the fractions containing the sex pili were subjected to isoelectric focusing. About 5 mg of intact pili of approximately 98% purity were obtained by this method from about 100 g (wet weight) of cells. 7.0.Assay for sex pili. The assay for sex pili was modified from that of Ippen and Valentine (9). 35Slabeled MS2 phage was prepared in a vigorously aerated flask at 37 C, containing 1 liter of IC broth, 814 on August 5, 2020 by guest

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Functions of F Pili in Mating-Pair Formation and Male Bacteriophage Infection Studied by Blending Spectra and Reappearance Kinetics

Cited by 25 publications

References 21 publications

Early Stages of Conjugation in Escherichia coli

Early Stages of Conjugation in Escherichia coli

Effects of high temperature on Escherichia coli F pili

Purification of Sex Pili from Escherichia coli Carrying a Derepressed F-Like R Factor

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