In 1988, John Cairns, Julie Overbaugh and Stefan Miller published a paper suggesting that some mutations in E. coli could arise in a Lamarckian fashion in response to the presence of a particular compound.
Their paper was controversial, to say the least. However, the phenomenon that they observed spurred much research that has led to a new understanding of mutagenesis and the dynamic nature of the stationary phase of cell growth.
The original experiments were carried out in E. coli D lac pro F' lacZam.
Similar results were later found in other strains of E. coli and in Salmonella typhimurium, Bacillus subtilis, Pseudomonas sp., and Clostridium sp.
Much of the recent work has been carried out in E. coli FC40 which carried a deletion of the lac region on the chromosome and an F' lac- episome in which a +1 frameshift mutation in lacZ is responsible for the Lac- phenotype.
The phenomenon is illustrated below:
In this experiment, E. coli FC40 is plated on minimal media plates containing lactose. After 2 days the number of Lac+ colonies increases dramatically (circles). However, the number of cells on the plate (triangles) does not.
Key features of this phenomenon are:
- Mutations arise among non-dividing cells after selective pressure is applied.
- The selective agent (lactose in the above example) is required.
- Mutations that are not selected do not appear.
Characterization of these mutants, however, revealed that
- The revertants were almost exclusively -1 frameshifts.
- Recombination functions were required for mutants to appear.
- Mutatnts were only observed at a high rate if the lac operon was carried on the episome and if conjugation functions were expressed.
The fact that the revertants were nearly all -1 frameshifts and
that they arose at a high rate suggested that an error-prone
DNA synthesis was involved rather than a random mutagenesis event.
We now know that three DNA polymerases (PolII,
PolIV and PolV)
are induced in stationary phase bacteria. PolII
(encoded by polB)
and PolIV (encoded by dinB) are relevant here.
PolII has a low error rate and normally reinitiates DNA synthesis downstream of gaps. PolIV is an error-prone DNA polymerase. In stationary phase cells, the two enzymes are in competition with one another. PolIV is thought to be responsible for 50% of the adaptive mutations that are observed.
The observations that recombination as well as conjugation functions are involved in the appearance of adaptive mutations can be understood by recalling that F plasmids encode their own transfer proteins and that these can and will be expressed in stationary phase cells. One of these functions, TraI, normally nicks the F plasmid as the first step in initiating transfer of a single DNA strand from the donor cell to the recipient.
However, this free end can also serve as a substrate for recombination - RecA mediated strand exchange could occur with another F plasmid (such as might exist in the aftermath of F plasmid replication). If so, then a process analogous to postreplicative repair but using an error-prone DNA polymerase could give rise to the adaptive mutations.
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|
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| DNA polymerase |
|
| PolI | RNA primer removal |
| PolII | Error-free re-initiation of DNA synthesis at short gaps or nicks |
| PolIII | The replicative polymerase |
| PolIV | Error-prone, stress induced & stationary phase enzyme. Introduces frame-shifts. |
| PolV | Error-prone, SOS induced enzyme. Copies over lesions such as thymine dimers. |
| Adaptive
mutation: implications for evolution Patricia L. Foster Abstract PDF Full Text (Size: 163K) BioEssays 22 (12): 1067 - 1074 (2000) |
