New insights into the bacterial immune system

MksBEFG defense system is widespread. (A) Representation of the mksFEBG operon organization from different organisms: (i) C. glutamicum ATCC13032, (ii) M. smegmatis mc2 155, (iii) P. aeruginosa UCBPP-PA14 and (iv) P. putida KT2440. (B) Plasmid copy number of low copy (pBHK18) and high copy number plasmids (pJC1) relative to oriC number per cell, analyzed by qPCR. Ratios were compared between C. glutamicum WT (MB001), ΔmksB, ΔmksG, ΔmksF cells grown in BHI medium with selection antibiotic (mean ± SD, n = 3). Plasmids pBHK18 and pJC1 were extracted from C. glutamicum WT, ΔmksB, ΔmksG, ΔmksF cells grown in BHI medium with antibiotic selection, visualization of extracted DNA on 0.8% agarose gels. (C) Phylogenetic analysis of MksG-like proteins (organized into DUF3322 and DUF2220 domains) using the SMART platform reveals the distribution between Gram-negative and Gram-positive bacteria and archaea. Credit: Nucleic acid research (2023). DOI: 10.1093/nar/gkad130

A research team from Kiel University describes an unknown defense mechanism in bacteria that selectively fends off foreign and potentially harmful genetic information.

Since the coronavirus pandemic, the particularly rapid evolutionary adaptability of microorganisms such as bacteria or viruses has been brought into the public spotlight. For example, when viruses develop the ability to infect new host organisms or bacteria develop antibiotic resistance, the uptake of new genetic information from other microorganisms allows them to rapidly express evolutionarily advantageous traits.

Bacteria, for example, take up foreign DNA through a process called horizontal gene transfer, which is much faster than vertical inheritance from generation to generation.

But any living organism also faces risks by taking in foreign genetic information, as it can potentially be dangerous if, for example, important genes are damaged by integration into its own chromosome, causing major disadvantages for the organism as a whole. Therefore, bacteria have evolved several mechanisms that protect them from absorbing harmful DNA. Many of the molecular processes involved were discovered in recent years, leading to the recent coining of the term “bacterial immune system.”

Now a team from the Microbial Biochemistry and Cell Biology Group at the Department of General Microbiology at Kiel University has elucidated the function of a new defense mechanism that can identify and, if necessary, degrade certain independent and mobile DNA structures called plasmids in bacteria cells – while there distinguish between useful and harmful genetic information.

Using the bacterium Corynebacterium glutamicum as an example, the researchers showed that the so-called Mks protein system has an additional element that can bind to plasmid DNA and cut it apart. The Kiel researchers led by Professor Marc Bramkamp published their new findings in Nucleic acid research.

Proteins for DNA organization can also defend against plasmids

Plasmids are small, usually circular, double-stranded DNA molecules that can replicate independently of the chromosome in their host cell. They play an important role in the ecology and evolution of bacteria, as they are an important vehicle for lateral gene transfer, enabling the rapid transfer of genetic information and thus the expression of selection advantages. In principle, all bacteria can exchange plasmids with each other, even across species.

This happens directly from bacterium to bacterium via a transfer mechanism known as conjugation. Both advantageous and disadvantageous plasmids use such bridges between bacterial cells to switch from one bacterium to another.

“How the bacterial organism deals with foreign DNA from newly transferred plasmids has been little researched so far,” Manuela Weiß, Ph.D. points out students in Bramkamp’s research group. “In previous research, we have investigated systems that are generally involved in the organization of DNA in bacterial cells and, among other things, ensure the packing of genetic information into the compressed form of chromosomes,” continues Weiß.

In this context, the research team obtained initial indications that C. glutamicum possesses two such systems, one of which is not involved in the organization of the chromosome, but can prevent the multiplication of certain plasmids, although the mechanism responsible for this was previously unknown.

Now the Kiel researchers, together with experts led by Dr. Anne Marie Wehenkel from the Institut Pasteur in Paris discovered the DNA scissors of the Mks system in a structural study. “We were able to prove experimentally that this new subunit of the Mks system forms a specific protein, a so-called nuclease, which can cut DNA. This element is tasked with degrading plasmids to keep harmful DNA away from the bacterial cell, while the other components of the Mks system are important for the recognition of plasmid DNA,” says Weiß.

To distinguish between useful and harmful plasmids

The researchers then followed up on the observation that the Mks system apparently only degrades certain plasmids, and that it must therefore be linked to a selection mechanism. An important advantage here is that Bramkamp’s research group works with the bacterium C. glutamicum, an organism that naturally possesses this system. Its functions can therefore be studied in vivo without changing its cell biological properties by transferring it to a model system.

“Bacteria use certain plasmids as a source of new, not immediately vital, genetic information. It is therefore obvious that a defense mechanism must be selective and not destroy all plasmids,” says Bramkamp.

“We were able to prove that in C. glutamicum there is in fact a directed selection for beneficial and harmful genetic information. When we artificially switched off the Mks system and thus all plasmids remained in the bacterial cells, there were harmful effects on the cell, possibly triggered by DNA stress, were evident. However, these did not occur when the defense mechanism was active,” continues Bramkamp.

With the current work, the Kiel researchers present important new findings about the bacterial immune system in general, which expand the understanding of plasmids as mediators of not only beneficial but also harmful genetic information. In the future, they will investigate which molecular mechanisms enable bacterial cells to distinguish between “good” and “bad” mobile DNA.

The new results are not only important for the general understanding of the organization and reproduction of bacterial life. The increasingly precise study of the bacterial immune system could also help to better meet applied challenges – and, for example, better model and predict the development of antibiotic resistance in certain bacterial populations in the future.

More information:
Manuela Weiß et al., The MksG nuclease is the executor of the bacterial plasmid defense system MksBEFG, Nucleic acid research (2023). DOI: 10.1093/nar/gkad130

Journal information:
Nucleic acid research

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