No new proteins in Lenski's experiment
Long Term Evolutionary Experiment conducted on Escherichia coli populations by Richard Lenski and his coworkers (also known as Lenski's experiment) is frequently cited in defense of evolutionary theory (for example by showing that fitness increased in these populations over time). Because of that many creationists are well aware of it and have already made quite a lot of claims regarding this experiment (some wilder than the others). That is exactly the reason why I am making this blog post - to respond to a claim regarding "no newly evolved proteins" by Escherichia populations. It is frequently made alongside the claim that "organisms in the experiment are still just bacteria even after tens of thousands of generations".
This is a great example of what I've talked about in my very first article on this blog Creationism is not scientifically valid - why I've created this blog? What I mean specifically is making baseless claims that supposedly prove evolution wrong and treating them as obviously proven facts while showing not even remotely any evidence in support of them. You see, the claim that Lenski's bacteria still being bacteria assumes that bacteria should completely rewire their biology and evolve into a completely new domain of life during the span of the experiment. The problem is that molecular data shows that species of Escherichia diverged from each other around tens of millions of years ago (depending on which species we take a look at). Simply put - most of lineages from genus Escherichia are still members of that genus (aside from Shigella lineages) after much longer than LTEE was running. If it comes to closest relatives of Escherichia (aside from Shigella nested within E.coli), specifically genus Salmonella, they have diverged from each other between 100 and 160 million years ago and yet they're still members of the same family (Enterobacteriaceae). Similarly, eukaryotes (plants, animals, fungi etc.) diverged from prokaryotes (Bacteria and Archaea) around 2 billion years ago. This means that bacteria probably "stayed bacteria" for BILLIONS of years. In other words - creationists demand something that would take millions and millions of years to happen in a mere couple of decades. Calling this notion ludicrous is an incredible understatement.
The claim about "no new proteins" is, as you might expect, completely baseless as well and rests on the "trust me bro" logic. But this claim is laughable not only due to this reason. In actuality, it is even more hilarious since the core claim that new proteins did not evolve in Lenski's experiment is... not even true! And I am here to prove it.
Genomic structure of amplified segments in Cit+ lineage with visible production of novel Rnk-CitG 89 aa Fusion Protein
First example of novel protein is related to the acquisition of Cit+ trait in population Ara-3. The Cit+ trait refers to an ability to grow on citrate in the presence of oxygen. E. coli normally dooesn't have that ability (including the ancestors of Lenski's experimental populations). The way this feature originated in the experiment was through the duplication of a genomic region which placed a copy of citrate-succinate antiporter gene under the control of a copy of promoter sequence from another gene. However, this mutation was not incredibly precise and because of that, an additional part of rnk gene's coding sequence was placed alongside an additional part of coding sequence of citG gene. This fusion gene was also placed under the control of the duplicated promoter. As a result, the Cit+ bacteria from this specific experiment began the synthesis of not only the citrate-succinate antiporter but also of the novel fusion protein named by researchers as Rnk-CitG 89 aa Fusion Protein. I have to also point out that as far as I'm aware this protein has no known function and it is certainly not necessary for the functioning of the Cit+ trait. That being said it may have some unknown function and it is certainly a new protein as the fusion gene encoding it did not exist in the ancestral strain.
A large deletion placed an ancestrally non-transcribed region (colored orange in section A) downstream of a strong promoter which led to new transcription (synthesis of mRNA) and translation (translating that mRNA into protein).
The second instance of production of a protein not present in LTEE ancestor is related to de novo gene birth. One population (specifically Ara-6) experienced a large deletion which "cut out" a couple of genes. This brought together a strong promoter and a previously non-coding sequence which led to the synthesis of a protein of a length of 117 amino acids. Now, some of creationists may become excited and try to point out that "THIS IS CLEARLY A DEGRADATION". In a sense, yes, but such "degradation" wouldn't be necessary as the placement of a strong promoter in front of a non-coding sequence could be achieved via other mechanisms such as duplication and illegitimate recombination (joining together completely unrelated DNA sequences) could place such a promoter copy without any deletions. Such effect could also be accomplished via the insertion of promoters from insertion sequences and further "strengthening" of such promoters. Insertion of promoters from insertion sequences has been documented in the same study regarding de novo 117 amino acid-long protein and it was shown that these promoters are not effective enough as to support reliable translation (even though this isn't always the case). Because of that such promoters would have to evolve the ability for stable translation in the first place which is supported empirically as non-coding regions have repeatedly shown the ability to evolve into stable promoters. Due to this phenomenon it would be possible as well to gain a new promoter via accumulation of single nucleotide polymorphisms (SNPs) and single nucleotide insertions upstream of a specific sequence since this mechanism could also allow the formation of DNA motifs necessary for the emergence of promoters. Additionally, novel proteins don't always have to stem from sequences homologous to other protein-coding ones as a promoter may be inserted into or independently evolve near any non-coding sequence and allow its expression.
Coming back to the de novo protein, we can learn an interesting thing from genomic analyses. This de novo protein is homologous (shows sequence similarity) to hypothetical proteins (proteins predicted to exist within a given genome but for the production of which there is no evidence so far) in other bacteria. This suggests that similar proteins either evolved independently in other bacteria or have been an ancestral feature which has been lost in LTEE's ancestor, just to be evolved independently in the Ara-6 population. This means that this protein may not be entirely new in the biological world. Regardless of the source of origin this is still an example of a new protein since there is no other known organism producing this exact protein and it certainly was not produced in the ancestor of bacterial populations from Lenski's experiment. So far, unfortunately, it wasn't determined if this protein has any function.
The study on de novo gene birth in Lenski's experiment also provides an interesting insight into the origin of new genes and proteins in general. While some proto-genes originated as early as 4000 generations into the experiment, the estimated average rate of formation of new proto-genes has been estimated to be around 1 per 60 000 generations. This is a substantial portion of the experiment's generations (as it is currently at around 80 000 generations). This proves that the emergence of new genes and proteins does not constantly occur constantly not because it is impossible for evolution but rather that such events don't occur that frequently and take a substantial amount of time. It is important to keep in mind that this is not to be used as a 1:1 comparison on the rate of emergence of de novo genes as various organisms feature substantial genomic differences. Aside from time limitations the amount of currently evolved novel proteins may stem from the impact of novel proteins on the fitness of bacterial populations in this experiment. If a novel protein evolves but its presence is detrimental it will probably sooner or later be outselected from the population and if its presence is neutral it may disappear from the population due to genetic drift (random population changes) before it becomes beneficial.
After reading this article, some creationists will probably claim something similar to "these are not TRULY new proteins because genes coding them stem from pre-existing information!". To that I would say that the criterion of "TRULY new" was not specified in the first place while examples I gave would be plainly understood as new and thus such a tactic can be considered a form of moving the goalpost. Furthermore, evolution has been shown to largely rely on duplication and modification of already existing information - a thing I pointed out in my first post. Nevertheless, the incident related to the new proteins in the Lenski's experiment clearly shows that new proteins that were not synthesized in a specific ancestor can later evolve in their descendants and it also shows that creationists clearly don't care about a robust amount of evidence found in scientific literature.
After reading this article, some creationists will probably claim something similar to "these are not TRULY new proteins because genes coding them stem from pre-existing information!". To that I would say that the criterion of "TRULY new" was not specified in the first place while examples I gave would be plainly understood as new and thus such a tactic can be considered a form of moving the goalpost. Furthermore, evolution has been shown to largely rely on duplication and modification of already existing information - a thing I pointed out in my first post. Nevertheless, the incident related to the new proteins in the Lenski's experiment clearly shows that new proteins that were not synthesized in a specific ancestor can later evolve in their descendants and it also shows that creationists clearly don't care about a robust amount of evidence found in scientific literature.
This is a picture of a "smiling" neutrophil.
Sources:
1. E.coli long-term evolution experiment: https://en.wikipedia.org/wiki/E._coli_long-term_evolution_experiment
2. Bacteria: https://en.wikipedia.org/wiki/Bacteria
3. Cryptic Lineages of the Genus Escherichia: https://pmc.ncbi.nlm.nih.gov/articles/PMC2765150/
4. Shigella: https://en.wikipedia.org/wiki/Shigella
5. Molecular evolution of Salmonella enterica serovar Typhimurium and pathogenic Escherichia coli: From pathogenesis to therapeutics: https://www.sciencedirect.com/science/article/abs/pii/S1567134807001748
6. Enterobacteriaceae: https://en.wikipedia.org/wiki/Enterobacteriaceae
7. Determining Divergence Times of the Major Kingdoms of Living Organisms with a Protein Clock: https://www.science.org/doi/abs/10.1126/science.271.5248.470
8. Genomic Analysis of a Key Innovation in an Experimental E. coli Population: https://pmc.ncbi.nlm.nih.gov/articles/PMC3461117/
9. Promoter recruitment drives the emergence of proto-genes in a long-term evolution experiment with Escherichia coli: https://pmc.ncbi.nlm.nih.gov/articles/PMC11101190/
10. Gene duplication: https://en.wikipedia.org/wiki/Gene_duplication
11. An Appraisal of the Potential for Illegitimate Recombination in Bacterial Genomes and Its Consequences: From Duplications to Genome Reduction: https://pmc.ncbi.nlm.nih.gov/articles/PMC403640/
12. Illegitimate recombination: https://en.wikipedia.org/wiki/Illegitimate_recombination
13. Random sequences rapidly evolve into de novo promoters: https://www.nature.com/articles/s41467-018-04026-w
14. Hypothetical protein: https://en.wikipedia.org/wiki/Hypothetical_protein
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