Modulating Campylobacter jejuni virulence by changing its rRNA methylation pattern.

Significance 

The ribosomal RNA molecules (rRNAs) of all bacteria are modified with an array of methyl groups, which serve to fine-tune the process of protein synthesis. These modifications are added to specific rRNA nucleotides by methyltransferase enzymes, and one such enzyme, TlyA, is found in several pathogenic bacteria. The TlyA ortholog in Campylobacter jejuni (termed TlyAI) adds a methyl group to a single specific rRNA nucleotide in the large subunit of the ribosome, while the ortholog of Mycobacterium spp. (termed TlyAII) methylates this same site plus an additional rRNA nucleotide in the small ribosomal subunit. Loss of the respective TlyA enzymes in Mycobacterium tuberculosis and C. jejuni alters their sensitivity to the antibiotic capreomycin and reduces an array of their virulence properties. The change in antibiotic sensitivity fits well with our understanding of ribosome function, because the methylations are situated on nucleotides within the capreomycin binding site. However, the mechanism by which the rRNA methylations affect virulence in the absence of antibiotics has remained unclear.

Dr. Agnieszka Sałamaszyńska-Guz and Dr. Izabela Serafińska from Warsaw University of Life Sciences together with Dr. Paweł Bącal at University of Warsaw and Professor Stephen Douthwaite at University of Southern Denmark have investigated these effects in C. jejuni. They showed that inactivation of the tlyAI gene in C. jejuni reduces several of the pathogen’s virulence traits including its ability to form biofilms and to adhere to and invade human intestinal epithelial cells. These traits can be to a large extent recovered by engineering an active copy of the  tlyAI gene back into the C. jejuni null-mutant at a different site in its chromosome.

In their most recent work published in Cellular Microbiology, the research team extended the investigation, this time using the mycobacterial tlyAII gene for complementation of the C. jejuni null-mutant. When expressed in C. jejuni, the mycobacterial TlyAII enzyme methylates the same two rRNA nucleotides as it would in its original Mycobacterium host. Addition of the two methyl groups not only restores the ability of C. jejuni to act as a pathogen, but surprisingly enhances some (but not all) of its virulence traits. For instance, the biofilms formed by the C. jejuni tlyAII-recombinant were more uniform and significantly denser, surpassing those of the wild-type. Furthermore, it was shown that the adhesion and invasion capacity of C. jejuni on Caco‐2 human colon epithelial cells, which was reduced to less than half that of the wild‐type following the loss of TlyAI‐directed methylation, was markedly increased upon complementation with the mycobacterial tlyAII gene. In contrast, other traits such as motility, survival time in epithelial cells, and sensitivity to capreomycin-type antibiotics were not significantly changed compared to C. jejuni cells expressing the original tlyAI gene.

In a nutshell, the authors demonstrate that the different methylation patterns on the C. jejuni rRNA affect its ability to be an effective pathogen. The authors propose that the methylations subtly modulate the rRNA’s function during the translation of certain mRNAs and thereby change the relative proportions of proteins in the bacterial cell. Such changes in the bacterial proteome would consequently alter the phenotype and virulence properties. These researchers are currently mapping the proteomes of the various C. jejuni recombinants using mass spectrometric approaches to identify key proteins involved in pathogenicity. It is envisaged that this approach will lead to a better understanding of virulence mechanisms and form a basis for finding novel means of treating Campylobacter jejuni infections and, ultimately, infections with other bacterial pathogens.

Modulating Campylobacter jejuni virulence by changing its rRNA methylation pattern. - Medicine Innovates

About the author

Agnieszka Sałamaszyńska-Guz is currently employed as an assistant professor at the Warsaw University of Life Sciences, working within the Division of Microbiology at the Institute for Veterinary Medicine. She obtained her PhD in 2006 working on bacterial toxins/hemolysins. During the last two decades, Dr Sałamaszyńska-Guz’s research has focused on elucidating the virulence mechanisms of Campylobacter species. These bacterial pathogens are the major cause of food poisoning worldwide, and the main agents in gastroenteritis and related intestinal disorders in Europe and North America. The most widespread of these species, Campylobacter jejuni, colonizes the human intestine by forming biofilms that facilitate attachment to and invasion of epithelial cells. Recently, Dr Sałamaszyńska-Guz received a prestigious grant from the Polish National Science Centre [2018/30/M/NZ6/00429] to investigated these processes.

She has shown that C. jejuni virulence can be attenuated by changing single modifications on the pathogen’s ribosomes, the structures where proteinaceous virulence factors are synthesized. Working in collaboration with the University of Southern Denmark, her mass spectrometric analyses of the proteomes of these attenuated strains revealed that that a discreet number of specific virulence factors are required to cause human disease. The identification of such factors presents unique targets for therapeutic intervention. Dr Sałamaszyńska-Guz’s immediate focus is to help reduce the incidence of Campylobacter colonization in commercial poultry (the main source of this pathogen) and to limit subsequent human infections. Longer-term aims include applying this approach to other bacterial pathogens, in particular those resistant to current clinical antibiotics.

Reference

Sałamaszyńska-Guz A, Serafińska I, Bącal P, Douthwaite S. Virulence properties of Campylobacter jejuni are enhanced by displaying a mycobacterial TlyA methylation pattern in its rRNA. Cell Microbiol. 2020 Jul;22(7):e13199.

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