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Examples of Beneficial Mutations and Natural Selection
from The Evolution Evidence Page
Scientists have shown that beneficial mutations do occur to produce brand new alleles (variants of genes) that improve an organism's chances of survival in a particular environment. Natural selection has been demonstrated to increase the frequency of these alleles in a population. The easiest way to demonstrate this is from experiments based upon lines of organisms developed from clones (genetically identical by definition) of a single individual. The following examples are from pages 183 - 186 the book "Selection - The Mechanism of Evolution" by Graham Bell, a book which I recommend highly to anyone who wants to learn more about evolution, especially if they like to see good evidence for the claims made for natural selection. I list the original references for some of these so that anyone who wants more details will know where to look. The title of the section from which these examples come is "A Single Clone of Microbes Will Readily Respond to Natural Selection". I start out with some examples of simple mutations, but in the section on evolution of new metabolic pathways, I present examples of series of mutations that "build on one another" to add new metabolic pathways for the organism. This is the very type of "new genetic information" that creationists frequently claim is impossible.
1.) Adaptation to High and Low Temperatures by E. coli.
A single clone of E. coli was cultured at 37 C (that is 37 degrees Celsius) for 2000 generations. A single clone was then extracted from this population and divided into replicates that were then cultured at either 32 C , 37 C, or 42 C for a total of another 2000 generations. Adaptation of the new lines was periodically measured by competing these selection lines against the ancestor population. By the end of the experiment, the lines cultured at 32 C were shown to be 10% fitter that the ancestor population (at 32 C), and the line cultured at 42 C was shown to be 20% more fit than the ancestor population. The replicate line that was cultured at 37 C showed little improvement over the ancestral line.
Bennett, A.F., Lenski, R.E., & Mittler, J.E. (1992). Evolutionary adaptation to temperature I. Fitness responses of Escherichia coli to changes in its thermal environment. Evolution, 46:16-30.
2.) Adaptation to Growth in the Dark by Chlamydomonas.
Chlamydomonas is a unicellular green algae capable of photosynthesis in light, but also somewhat capable of growth in the dark by using acetate as a carbon source. Graham Bell cultured several clonal lines of Chlamydomonas in the dark for several hundred generations. Some of the lines grew well in the dark, but other lines were almost unable to grow at all. The poor growth lines improved throughout the course of the experiment until by 600 generations they were well adapted to growth in the dark. This experiment showed that new, beneficial mutations are capable of quickly (in hundreds of generations) adapting an organism that almost required light for survival to growth in the complete absence of light.
3.) Selection for Large Size in Chlamydomomas
Bell also selected clonal lines of Chlamydomonas for size by passing cultures through a fine filter and discarding the cells that were not retained on the filter. He reports that although this method was not very effective at retaining the largest cells (due to inconsistencies in the filter pore size), after forty generations of this selection technique, the cell diameter had increased by an average of about 1 phenotypic standard deviation.
4.) Adaptation to a Low Phosphate Chemostat Environment by a Clonal Line of Yeast
P.E. Hansche and J.C. Francis set up chemosats to allow evolution of a single clonal line of beer yeast in a phosphate limited (due to high pH) environment. (A chemostat is a device that allows the propagation of microorganisms in an extremely constant environment.) The yeast clones grew slowly for about the first 180 generations when there was an abrupt increase in population density. This was later shown to be due to better assimilation of the phosphate, presumably due to an improvement in the permease molecule. (Permease is an enzyme that controls what is allowed to come into the cell through the yeast's cell membrane.) After about 400 generations, a second improvement in cell growth rates occurred because of a mutation to the yeast's phosphatase (an enzyme that improves the cells ability to use phosphate). The phosphatase became more active overall, and its optimal pH (the pH where it is most active) was raised. Finally, a third mutant appeared after 800 generations that caused the yeast cells to clump. This raised the population density in the chemostat because individual cells were no longer being washed out of chemostat (which is one of the methods that the chemostat uses to maintain very uniform conditions) as quickly as they had prior to the mutation. (This is just speculation on my part, but I wonder if it wasn't under some similar conditions that multi-cellularity became favored over unicellularity - perhaps on a sea bed or river bottom.)
This experiment was repeated, and the same mutations occurred, but in different orders. Also, in one replication, the processing of phosphate was improved by a duplication of the gene that produces phosphatase. This is experimental evidence of an extremely important mechanism in evolutionary history! It is also a particularly elegant experiment because not only was all of this adaptation shown to occur in clonal lines (descended from a single individual), but the authors also determined the exact mutations that caused the improved adaptations by sequencing the genes and proteins involved.
Francis, J.E., & Hansche, P.E. (1972) Directed evolution of metabolic pathways in microbial populations. I. Modification of the acid phosphatase pH optimum in Saccharaomyces cervisiae. Genetics, 70: 59-73.
Francis, J.E., & Hansche, P.E. (1973) Directed evolution of metabolic pathways in microbial populations. II. A repeatable adaptation in Saccharaomyces cervisiae. Genetics, 74:259-265.
Hansche, P.E. (1975) Gene duplication as a mechanism of genetic adaptation in Saccharaomyces cervisiae. Genetics, 79: 661-674.
These examples are not from Bell's book:
5.) Evidence of genetic divergence and beneficial mutations in bacteria after 10,000 generations
Papadopoulos, D., Schneider, D., Meier-Eiss, J., Arber, W., Lenski, R. E., Blot, M. (1999). Genomic evolution during a 10,000-generation
experiment with bacteria. Proc. Natl. Acad. Sci. U. S. A. 96: 3807-3812
Edited by John R. Roth, University of Utah, Salt Lake City, UT, and approved February 3, 1999 (received for review July 21, 1998)
Molecular methods are used widely to measure genetic diversity within populations and determine relationships among species. However, it is difficult to observe genomic evolution in action because these dynamics are too slow in most organisms. To overcome this limitation, we sampled genomes from populations of Escherichia coli evolving in the laboratory for 10,000 generations. We analyzed the genomes for restriction fragment length polymorphisms (RFLP) using seven insertion sequences (IS) as probes; most polymorphisms detected by this approach reflect rearrangements (including transpositions) rather than point mutations. The evolving genomes became increasingly different from their ancestor over time. Moreover, tremendous diversity accumulated within each population, such that almost every individual had a different genetic fingerprint after 10,000 generations. As has been often suggested, but not previously shown by experiment, the rates of phenotypic and genomic change were discordant, both across replicate populations and over time within a population. Certain pivotal mutations were shared by all descendants in a population, and these are candidates for beneficial mutations, which are rare and difficult to find. More generally, these data show that the genome is highly dynamic even over a time scale that is, from an evolutionary perspective, very brief.
6.) Adaptation of yeast to a glucose limited environment via gene duplications and natural selection
When microbes evolve in a continuous, nutrient-limited environment, natural selection can be predicted to favor genetic changes that give cells greater access to limiting substrate. We analyzed a population of baker's yeast that underwent 450 generations of glucose-limited growth. Relative to the strain used as the inoculum, the predominant cell type at the end of this experiment sustains growth at significantly lower steady-state glucose concentrations and demonstrates markedly enhanced cell yield per mole glucose, significantly enhanced high-affinity glucose transport, and greater relative fitness in pairwise competition. These changes are correlated with increased levels of mRNA hybridizing to probe generated from the hexose transport locus HXT6. Further analysis of the evolved strain reveals the existence of multiple tandem duplications involving two highly similar, high-affinity hexose transport loci, HXT6 and HXT7. Selection appears to have favored changes that result in the formation of more than three chimeric genes derived from the upstream promoter of the HXT7 gene and the coding sequence of HXT6. We propose a genetic mechanism to account for these changes and speculate as to their adaptive significance in the context of gene duplication as a common response of microorganisms to nutrient limitation.
Brown CJ, Todd KM, Rosenzweig RF (1998) Multiple duplications of yeast hexose transport genes in response to selection in a glucose-limited environment. Mol Biol Evol 1998 Aug;15(8):931-42 Nature 387, 708 - 713 (1997)
7.) Molecular evidence for an ancient duplication of the entire yeast genome
KENNETH H. WOLFE AND DENIS C. SHIELDS
Gene duplication is an important source of evolutionary novelty. Most duplications are of just a single gene, but Ohno proposed that whole-genome duplication (polyploidy) is an important evolutionary mechanism. Many duplicate genes have been found in Saccharomyces cerevisiae, and these often seem to be phenotypically redundant. Here we show that the arrangement of duplicated genes in the S. cerevisiae genome is consistent with Ohno's hypothesis. We propose a model in which this species is a degenerate tetraploid resulting from a whole-genome duplication that occurred after the divergence of Saccharomyces from Kluyveromyces. Only a small fraction of the genes were subsequently retained in duplicate (most were deleted), and gene order was rearranged by many reciprocal translocations between chromosomes. Protein pairs derived from this duplication event make up 13% of all yeast proteins, and include pairs of transcription factors, protein kinases, myosins, cyclins and pheromones. Tetraploidy may have facilitated the evolution of anaerobic fermentation in Saccharomyces.
8.) Evolution of a new enzymatic function by recombination within a gene.
Hall BG, Zuzel T
Proc Natl Acad Sci U S A 1980 Jun 77:6 3529-33
Mutations that alter the ebgA gene so that the evolved beta-galactosidase (ebg) enzyme of Escherichia coli can hydrolyze lactose fall into two classes: class I mutants use only lactose, whereas class II mutants use lactulose as well as lactose. Neither class uses galactosylarabinose effectively. In this paper we show that when both a class I and a class II mutation are present in the same ebgA gene, ebg enzyme acquires a specificity for galactosylarabinose. Although galactosylarbinose utilization can evolve as the consequence of sequential spontaneous mutations, it can also evolve via intragenic recombination in crosses between class I and class II ebgA+ mutant strains. We show that the sites for class I and class II mutations lie about 1 kilobase, or about a third of the gene, apart in ebgA. Implications of these findings with respect to the evolution of new metabolic functions discussed.
9.) Changes in the substrate specificities of an enzyme during directed evolution of new functions.
Biochemistry 1981 Jul 7 20:14 4042-9
Wild-type ebg enzyme, the second beta-galactosidase of Escherichia coli K12, does not permit growth on lactose. As part of a study of the evolution of new enzymatic functions, I have selected, from a lacZ deletion strain, a variety of spontaneous mutants that grow on lactose and other beta-galactoside sugars. Single point mutations in the structural gene ebgA alter the enzyme so that it hydrolyzes lactose or lactulose effectively; two mutations in ebgA permit galactosylarabinose hydrolysis, while three mutations are required for lactobionic acid hydrolysis. Wild-type ebg enzyme and 16 functional mutant ebg enzymes were purified and analyzed kinetically to determine how the substrate specificities had changed during the directed evolution of these new functions. The specificities for the biologically selected substrates generally increased by at least an order of magnitude via increased Vmax and decreased Km for the substrate. These changes were very specific for the selected substrate, often being accompanied by decreased specificities for other related substrates. The single, double, or triple substitutions in the enzymes did not detectably alter the thermal stability of ebg enzyme.
10.) 12% (3 out of 26) random mutations in a strain of bacteria improved fitness in a particular environment.
Contribution of individual random mutations to genotype-by-environment interactions in Escherichia coli
Susanna K. Remold* and Richard E. Lenski
Center for Microbial Ecology, Michigan State University, East Lansing, MI 48824
Edited by M. T. Clegg, University of California, Riverside, CA, and approved July 30, 2001 (received for review March 22, 2001)
Numerous studies have shown genotype-by-environment (G×E) interactions for traits related to organismal fitness. However, the genetic architecture of the interaction is usually unknown because these studies used genotypes that differ from one another by many unknown mutations. These mutations were also present as standing variation in populations and hence had been subject to prior selection. Based on such studies, it is therefore impossible to say what fraction of new, random mutations contributes to G×E interactions. In this study, we measured the fitness in four environments of 26 genotypes of Escherichia coli, each containing a single random insertion mutation. Fitness was measured relative to their common progenitor, which had evolved on glucose at 37°C for the preceding 10,000 generations. The four assay environments differed in limiting resource and temperature (glucose, 28°C; maltose, 28°C; glucose, 37°C; and maltose, 37°C). A highly significant interaction between mutation and resource was found. In contrast, there was no interaction involving temperature. The resource interaction reflected much higher among mutation variation for fitness in maltose than in glucose. At least 11 mutations (42%) contributed to this G×E interaction through their differential fitness effects across resources. Beneficial mutations are generally thought to be rare but, surprisingly, at least three mutations (12%) significantly improved fitness in maltose, a resource novel to the progenitor. More generally, our findings demonstrate that G×E interactions can be quite common, even for genotypes that differ by only one mutation and in environments differing by only a single factor.
Evolution of a Unicellular Organism into a Multicellular Species
Starting from single celled animals, each of which has the capability to reproduce there is no sex in the sense that we think of the term. Selective pressure has been observed to convert single-cellular forms into multicellular forms. A case was observed in which a single celled form changed to multicellularity.
Boxhorn, a student of Boraas,writes:
Coloniality in Chlorella vulgaris
Boraas (1983) reported the induction of multicellularity in a strain of Chlorella pyrenoidosa (since reclassified as C. vulgaris) by predation. He was growing the unicellular green alga in the first stage of a two stage continuous culture system as for food for a flagellate predator, Ochromonas sp., that was growing in the second stage. Due to the failure of a pump, flagellates washed back into the first stage. Within five days a colonial form of the Chlorella appeared. It rapidly came to dominate the culture. The colony size ranged from 4 cells to 32 cells. Eventually it stabilized at 8 cells. This colonial form has persisted in culture for about a decade. The new form has been keyed out using a number of algal taxonomic keys. They key out now as being in the genus Coelosphaerium, which is in a different family from Chlorella. "
Boraas, M. E. 1983. Predator induced evolution in chemostat culture. EOS. Transactions of the American Geophysical Union. 64:1102.
from Observed Instances of Speciation
Evolution of New Metabolic Pathways
Some of the experiments that I described previously could be said to have one weakness - they each only cover minor changes to an existing metabolic pathway. Pages 229 - 243 of Graham Bell's book ("Selection - The Mechanism of Evolution" Alexander Bell, 1997) also describe experiments that have demonstrated that organisms are capable of evolving whole new metabolic pathways, not just improving existing pathways. This is important to me because it shows that evolution is capable of producing something new, not just making minor improvements to existing pathways.
The most common experimental process that leads to evolution of a new pathway is this. An organism (usually bacteria) is put in a novel environment - an environment that contains some resource (chemical) that the organism has not been exposed to in the past. If that new chemical is the sole source of some chemical the organism requires for survival (e.g. carbon or nitrogen), most of the time, the organism will die. However, if any of the organisms' existing enzymes have the slightest ability to enhance reactions with the new resource, selection will strongly favor the duplication of the gene that produces that enzyme, and future mutations will improve the ability of the newly duplicated enzyme to process the new chemical resource. It's not very hard to see that the same factors that are manipulated in these experiments are going to occur in nature as well in the very long term. This duplication and divergence "strategy" (in quotes because evolution doesn't really have a strategy) of evolution appears to be capable of driving evolution from the first pre biotic self-replicators to the present incredibly complex and diverse life forms that occupy every imaginable niche of the planet earth.
1.) Modifying the fucose pathway to metabolize propanediol.
In normal anaerobic E. coli metabolism L-fucose is transported into the cell and converted into dihydroxyacetone phosphate (which is used for further metabolism) and lactaldehyde (which is a waste product). The lactaldehyde is then converted to propanediol which is actively excreted from the cell by a "facilitator" - a chemical that eases movement of another chemical through the cell membrane in either direction. When E. coli lines are exposed to an aerobic environment rich in propanediol, some individuals are able to utilize this former waste as a food source. This is made possible by a change to the enzyme that formerly converted the lactaldehyde to propanediol to reverse its action and convert the propanediol to lactaldehyde. The lactaldehyde can then be processed by the previously existing aerobic pathways that use lactaldehyde as a carbon and energy source.
Lin, E.C.C., & Wu, T.T. (1984) Functional divergence of the L-Fucose system in Escherichia coli. In R.P. Mortlock (ed.), "Microorganisms as Model Systems for Studying Evolution" (pp. 135-164) Plenum, New York.
2.) Exotic five-carbon sugars
Some five carbon sugars are very rare in nature, so very few organisms have the ability to use these exotic compounds in their metabolism. Robert Mortlock determined that the bacteria Klebsiella aerogenes was not immediately able to metabolize D-arabinose and xylitol by growing strains in media containing those compounds and noting the strains that were able to grow only after a lag time. This indicated that the original strain did not have the ability to process the compounds, but was able to evolve such a capability. Mortlock then went on to see how this capability was evolved.
In the case of D-arabinose, Mortlock showed that the arabinose could be utilized if it could be converted to D-ribulose by an enzyme (an isomerase). Unfortunately, K. aerogenes has no such isomerase for the conversion of D-arabinose. However, the isomerase for L-fucose has a low activity for D-arabinose. But, the bad news is that the L-fucose isomerase is normally produced only when the cell is exposed to fucose. Nonetheless, in a few individuals, mutations occurred that allowed the fucose isomerase to be produced at all times - not just when L-fucose is present. This is normally a bad thing and would be selected against because it wastes the cells resources by constantly producing an unneeded enzyme. In this situation though, the mutation is a very good thing, and allows the cell to survive because it can now metabolize arabinose (albeit rather poorly). Although production of the fucose isomerase has been deregulated, the structure of the isomerase itself has not been changed. The next mutation was a change to the isomerase to make it more effective in the conversion of arabitol to ribulose. Finally (although I can't tell from Bell's description if this was actually done in the experiments), the culture could be selected to regain control of the expression of the isomerase - so that it is produced only when arabitol is present.
Xylitol is also not normally metabolized, but Mortlock and his colleagues were able to develop strains (generally through spontaneous mutations, but sometimes with u.v. ray or chemical induced mutations) that could use it because ribitol dehydrogenase (which is usually present in the cells to convert ribitol to D-ribulose) was able to slightly speed up the conversion of xylitol to D-xylulose, for which metabolic pathways already exist. The ability of the strains to utilize xylitol was increased as much as 20 fold when first production of ribitol dehydrogenase was deregulated (the enzyme was produced all the time, not just when ribitol was present), then duplication of the ribitol dehydrogenase genes occurred, then the structure of the enzyme was changed such that its efficiency at working with xylitol was improved, and finally, in at least one case, a line regained control of the modified ribitol dehydrogenase gene so that the enzyme was only produced in the presence of xylitol. Here we have a complete example of a new metabolic pathway being developed through duplication and modification of an existing pathway.
Many papers were published concerning this group of experiments. For a review, see:
Hartley, B.S. (1984), Experimental evolution of ribitol dehydrogenase. In R.P. Mortlock (ed.), "Microorganisms as Model Systems for Studying Evolution" (pp. 23 - 54) Plenum, New York.
Bell goes on to give at least two more examples of the evolution of new metabolic pathways.
In his 1999 book "Finding Darwin's God", Kenneth Miller describes Barry Hall's series of experiments in which he deletes an important gene in a bacterium that allows them to use sugar as a food source. SeeParts is Parts, which is an excerpt from pg. 145 - 147 of "Finding Darwin's God to learn about this reseach.
5.) Adaptive Evolution Of Escherichia coli K-12 MG1655 On A Non-Native Carbon Source, L-1,2-Propanediol.
Appl Environ Microbiol. 2010 Apr 30. [Epub ahead of print]
Lee DH, Palsson BO.
Department of Bioengineering, University of California-San Diego, La Jolla, California 92093-0412.
Laboratory adaptive evolution can provide key information to address a wide range of issues in evolutionary biology. Such studies have been limited thus far by the inability to readily detect mutations on a genome-scale in evolved microbial strains. This limitation has now been overcome by recently-developed genome sequencing technology that readily allows us to identify all accumulated mutations that appear during laboratory adaptive evolution. In this study, we evolved Escherichia coli K-12 MG1655 on a non-native carbon source, L-1,2-propanediol (L-1,2-PDO) for approximately 700 generations. We found that; i) Experimental evolution of E. coli for approximately 700 generations in 1,2-PDO-supplemented minimal medium revealed the acquisition of the ability to use L-1,2-PDO as the sole carbon and energy source, from no growth at all initially, to a growth rate of 0.35 h(-1), ii) Six mutations detected by whole genome re-sequencing were accumulated in the evolved E. coli mutant over the course of adaptive evolution on L-1,2-PDO, iii) Five of six mutations were within coding regions, with IS5 insertion between two fuc regulons, iv) Two major mutations (mutations in fucO and its promoter) involved in L-1,2-PDO catabolism appeared early during adaptive evolution, v) Multiple defined knock-in mutant strains with all mutations had growth rates essentially matching that of the evolved strain. These results provide insight into the genetic basis underlying microbial evolution to growth on the non-native substrate. PMID: 20435762 [PubMed - as supplied by publisher]