The F Plasmid Is Transferred by Conjugation Between Bacteria

KEY CONCEPTS
-The free F plasmid is a replicon that is maintained at the level of one plasmid per bacterial chromosome.
-An F plasmid can integrate into the bacterial chromosome, in which case its own replication system is suppressed.
-The F plasmid encodes a DNA translocation complex and specific pili that form on the surface of the bacterium.
-An F-pilus enables an F-positive bacterium to contact an F-negative bacterium and to initiate conjugation.
Another example of a connection between replication and the propagation of a genetic unit is provided by bacterial conjugation, in which a plasmid genome or part of a host chromosome with an integrated episome is transferred from one bacterium to another.
Conjugation is mediated by the F plasmid, which is the classic example of an episome—an element that can exist as a free circular plasmid, or that can become integrated into the bacterial chromosome as a linear sequence (like a lysogenic bacteriophage). The F plasmid is a large, circular DNA approximately 100 kilobases (kb) in length.
The F plasmid can integrate at numerous sites in the E. coli chromosome, often by a recombination event involving certain sequences (called IS sequences) that are present on both the host chromosome and F plasmid. In its free (plasmid) form, the F plasmid utilizes its own replication origin (oriV) and control system, and is maintained at a level of one copy per bacterial chromosome. When it is integrated into the bacterial chromosome, this system is suppressed, and F DNA is replicated as a part of the chromosome.
The presence of the F plasmid, whether free or integrated, has important consequences for the host bacterium. Bacteria that are F-positive are able to conjugate (or mate) only with bacteria that are F-negative. Conjugation involves direct, physical contact between donor (F-positive) and recipient (F-negative) bacteria; contact is followed by one-way transfer of the F plasmid from the donor to the recipient (but never the other way). If the F plasmid exists as a free plasmid in the donor bacterium, it is transferred as a plasmid and the infective process converts the F-negative recipient into an F-positive state. If the F plasmid is present in an integrated form in the donor, the transfer process might also cause some or (rarely) all of the bacterial chromosome to be transferred.
Many plasmids have conjugation systems that operate in a generally similar manner, but the F plasmid was the first to be discovered and remains the paradigm for this type of genetic transfer.
A large (about 33 kb) region of the F plasmid called the transfer region is required for conjugation. It contains roughly 40 genes that are required for the transmission of DNA; FIGURE 1. summarizes their organization. The genes are arranged in loci named tra and trb. Most of them are expressed coordinately as part of a single polycistronic 32-kb transcription unit (the traY-I unit). traM and traJ are expressed separately. traJ is a regulator that turns on both traM and traY-I. On the opposite strand, finP is a regulator that codes for a small antisense RNA that turns off traJ.

Its activity requires expression of another gene, finO. Only four of the tra and trb genes, traD, traI, traM, and traY, in the major transcription unit are concerned directly with the transfer of DNA; most of these genes encode proteins that form a large membranespanning protein complex called a type 4 secretion system (T4SS).
These systems are common in bacteria, where they have been shown to be involved in the transport of various proteins and DNA across the bacterial cell envelope and are responsible for maintaining contacts between mating bacteria.

FIGURE 1. The tra region of the F plasmid contains the genes needed for bacterial conjugation.
F-positive bacteria possess surface appendages called pili (singular pilus) that are encoded by the F plasmid. The gene traA codes for the single subunit protein, pilin, that is polymerized into
the pilus extending from the inner to the outer membrane at the T4SS. At least 12 tra genes are required for the modification and assembly of pilin into the pilus and the stabilization of the T4SS.
The F-pili are hairlike structures, 2 to 3 μm long, that protrude from the bacterial surface. A typical F-positive cell has two to three pili. The pilin subunits are polymerized into a hollow cylinder, about 8 nm in diameter, with a 2-nm axial hole.
Mating is initiated when the tip of the F-pilus contacts the surface of the recipient cell. FIGURE 2. shows an example of E. coli cells beginning to mate. A donor cell does not contact other cells carrying the F plasmid, because the genes traS and traT encode “surface exclusion” proteins that make the cell a poor recipient in such contacts. This effectively restricts donor cells to mating with F-negative cells. (The presence of F-pili has secondary consequences; they provide the sites to which RNA phages and some single-stranded DNA phages attach, so F-positive bacteria are susceptible to infection by these phages, whereas F-negative bacteria are resistant.)

FIGURE 2. Mating bacteria are initially connected when donor F-pili contact the recipient bacterium.
Photo courtesy of Emeritus Professor Ron Skurray, School of Biological Sciences, University of Sydney.
The initial contact between donor and recipient cells is easily broken, but other tra genes act to stabilize the association; this brings the mating cells closer together. The F-pili are essential for initiating pairing, but retract or disassemble as part of the process by which the mating cells are brought into close contact. It is proposed that the T4SS provides the channel through which DNA is transferred. TraD is a so-called coupling protein encoded by F plasmids that is necessary for recruitment of plasmid DNA to the T4SS, and it may associate with the T4SS to be involved in the actual plasmid transfer.

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