Breaking down the microbiology world one bite at a time
Bacteria’s first response against antibiotics? Epigenetics
Microorganisms wrestle for control of spaces to colonize, as humans have throughout history. The first antibiotic humanity discovered, penicillin, was nothing other than one of the many molecules that microorganisms already used to fight each other. Since then, we have found and created more, but the truth is, we are late to the fight in evolutionary terms. Bacteria have developed many ways to adapt to antibiotics over millennia, and those ways give the bacteria antibiotic resistance.
The most common, or perhaps best-known cause of resistance, is the presence of antibiotic resistance genes, usually stored in small DNA rings called plasmids. Bacteria can copy these plasmids and pass them to their neighbors, allowing resistance to spread quickly. When exposed to antibiotics, these genes produce proteins that break down, expel, or modify the antibiotics into a harmless version of them. Bacterial DNA also mutates more often than eukaryotic DNA, and sometimes these mutations help them resist antibiotics. But all of these are slow changes, affecting the genetic information stored in DNA. There is also another, much faster way bacteria adapt, called epigenetics. Epigenetics deals with changes in DNA and RNA, small modifications that regulate the function of the larger molecules. Consider them legends or footnotes for the nucleic acids. But these tiny changes are mighty important.
Antibiotics act at a molecular level by stopping crucial functions of the cells, and one common way is to stop translation, the synthesis of proteins using an RNA template. This is done by a ribosome, a large molecular machine composed of ribosomal RNA (rRNA) and proteins. The ribosome takes the template RNA that has been transcribed from a gene, and assembles proteins by linking amino acids, the building blocks of proteins. Antibiotics like streptomycin, kasugamycin, and many others, work by blocking parts of the ribosome, impeding the synthesis of new proteins, which causes the bacteria to eventually die.
As early as 1984, researchers discovered that mutations in the genes encoding rRNA could make bacteria resistant to antibiotics, by slightly altering the ribosome so the drug can’t bind properly. In fact, epigenetic modifications of the rRNA are common, and improve ribosomal function. Researchers have now used a technology for RNA sequencing, called Nanopore, which allows them to know where the modifications are on the rRNA. The nucleic acids (DNA and RNA) go through a pore in a membrane, and each time a nucleotide goes through, it changes the electrical charge of the membrane slightly. This can be recorded and correlated. However, the small modifications on each nucleotide also generate a slight change, which can be recorded, but it is difficult to do so correctly.
With this technology, researchers tested what happened to the rRNA when bacteria cells were exposed to streptomycin and kasugamycin. The modifications on the rRNA of Escherichia coli cells treated with antibiotics were much fewer than in healthy bacteria. Further analysis confirmed that when exposed to the antibiotics, the bacteria produced new rRNA, and did not modify it. They also tested resistance to the antibiotics in normal E. coli and in E. coli mutants that did not have the proteins responsible for some of the rRNA modifications. The latter were much more resistant to the antibiotics, although still died to high concentrations of the drug.
Mutations in DNA happen over generations, but RNA modifications occur in minutes. Bacteria lacking the right gene or mutation will die in the presence of antibiotics, but in RNA modifications we have uncovered a new layer of protection that bacteria use to shield themselves from antibiotics. The trouble is that these modifications are not easy to measure and read, and there are many different types. Despite the new RNA sequencing techniques, the changes in the RNA are tiny, and we require specialized software and algorithms to make sense of the data correctly.
The researchers in this study have established one new such software, called NanoConsensus. This software uses four already-established algorithms to detect rRNA modifications, correcting for individual bias by checking with the other algorithms to see if the modifications are also picked up by them. NanoConsensus turned out to be much more precise with detecting the modifications than each individual algorithm.
Lastly, the researchers used NanoConsensus to check which modifications were important in E. coli resistance to streptomycin and kasugamycin. They found two key modifications of high importance. When E. coli mutants do not have the proteins that place these modifications, they are more resistant to the antibiotics.
To wrap it up, NanoConsensus and Nanopore RNA sequencing technology might allow us to detect how epigenetic resistance happens in other clinically relevant bacteria, and perhaps open the door to new treatments or combination therapies with drugs that block ribosomes with modified and unmodified rRNA, to improve survival and recovery in bacterial infections.
Link to the original post: Delgado-Tejedor, A., Medina, R., Begik, O. et al. Native RNA nanopore sequencing reveals antibiotic-induced loss of rRNA modifications in the A- and P-sites. Nat Commun 15, 10054 (2024).
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