The F factor (fertility factor) of Esch. coli was the first transfer factor to be discovered but in some ways it is not typical. It is unusual in that it can be inserted into the bacterial chromosome by a mechanism similar to that of the insertion of lambda (λ) phage into the chromosome and that it can then transfer chromosomal genes into recipient cells. This feature of its behavior dominated discussion of the nature of transfer factors for many years. The more recent discovery of non-transmissible plasmids and of many transfer factors that carry genes for drug resistance but seldom, if ever, become associated with the chromosome, has led to a broader understanding of the general nature of plasmids.
The F factor is a transfer factor that contains the basic genetic information for extrachromosomal existence and for self-transfer, but does not normally contain other identifiable genetic markers such as drug-resistance genes. Cells that contain the F plasmid free in the cytoplasm (F+ cells) have no unusual characters apart from the ability to produce pili and to transfer F to F- cells by conjugation. In a very small pro-portion of F+ cells F becomes inserted into the bacterial chromosome; such cells are able to transfer certain chromosomal genes into F- cells with high frequency and cultures of these cells are known as Hfr strains. The F factor is a small circular piece of double-stranded DNA.
When it becomes associated with the chromosome it may be inserted into it as a linear segment by a single recombination event. The F factor has not been shown to produce a specific integrase enzyme as does A phage integration of F is thought to occur simply by recombination between regions of genetic homology.
There are presumed to be several regions of homology between F and the chromosome of its normal host strain of Esch. coli as F may integrate at a number of different sites on the chromosome. When F is integrated into the chromosome in the Hfr state, it is replicated as part of the chromosome and inherited by the progeny cells in the same way that a prophage is inherited by lysogenized cells.
The transfer functions of F are not repressed in the chromosome; it still directs the formation of pili on the cell surface so that Hfr cells can conjugate and transfer DNA. In this state they transfer not only the F factor DNA but also part of the chromosome to which it is attached. During conjugation one strand of the plasmid DNA is opened and a linear piece of DNA is transferred into the recipient cell.
In this case a part of the F factor DNA is first transferred, followed by the part of the chromosome at one side of the site of insertion of F. As conjugation continues, more of the bacterial chromosome is pushed through into the recipient. However, the chromosome tends to break randomly during transfer so that, on average, only about 10 per cent is transferred. Only the rare cell that receives the whole chromosome will also receive the other portion of the F factor and become F.
All cells in a culture of a particular Hfr strain have F integrated at the same site in the chromosome. When Hfr cells conjugate with F- cells, the recipients acquire the donor cell chromosomal genes in a specific order—the order in which they occur on the chromosome. Mating cells can be broken apart mechanically by vigorous shaking so that the piece of DNA that is being transferred is broken.
If conjugation is interrupted at different times after the Hfr and F- cells have been mixed, the order of the genes on the donor cell chromosome can be mapped by determining the time when each gene first becomes detectable in the recipient cells. It is found that 120 minutes is required for the whole chromosome to be transferred.
Since there is almost perfect homology between the recipient cell chromosome and the transferred portion of chromosome, recombination occurs readily and there is quite a high probability that any introduced gene will be incorporated into the chromosome and inherited by the progeny. Hfr strains produce a high frequency of transfer of the early genes and thus a high frequency of recombination; hence the abbreviation Hfr.
Different Hfr strains have the F factor inserted at different sites. During conjugation, the genes are in the same order on the chromosome, but transfer starts at different points, and may proceed in either direction. Hfr strain may transfer genes in the order ABCDE . . . while another transfers PQRST . . . and a third transfers JIHGF . . . Such experiments, using a number of Hfr strains, revealed that the genetic map of Esch. coli was circular before it was technically possible to demonstrate the circular nature of the chromosome DNA.
The use of Hfr donors allows the study of much longer fragments of DNA than can be transferred between cells by transduction and it has made possible the construction of a circular map of the Esch. coli genome that locates the relative positions of some 300 genes. Transduction, by contrast, is particularly useful for analyzing the detailed arrangement of genes that are closely clustered on the chromosome.
In any culture of F+ cells i there is a small proportion of cells that have F integrated into the chromosome. Similarly, in any culture of Hfr cells there are a few in which F comes out of the chromosome again and reverts to the Free State. This is similar to the situation with A, prophage where, in a small proportion of cells, the prophage is excised from the chromosome and phage replication is resumed). As with λ phage, F is not always excised accurately and occasionally an F factor carries off some of the neighboring genes when it leaves the chromosome. An F factor that has picked up a portion of the chromosome in this way is known as an F-prime factor (F'). One well known example is the F-lac factor, which is an F' carrying the genes of the Esch. coli lac operon. When an F' factor transfers itself into another cell its associated genes function normally on the plasmid in the new host.
Thus, non-lactose-fermenting organisms become lactose-fermenters when they receive F-lac. When F -lac is transferred into Esch. Coli, the cells become diploid for the lac genes. The particular value of this partial diploidy is that it allows the study of interactions between different lac operon mutations within the same cell. This led to the discovery of the mechanism of control of inducible enzymes.