Bacterial Ribosome Heterogeneity by Truncated r-Proteins

Proteins are the building blocks of a cell. To generate a protein, the corresponding genetic information on the DNA is copied onto a temporary storage module, the RNA. An enormous complex called the ribosome reads the copied information on the RNA and connects different amino acids according to the genetic code to synthesize the desired protein. Ribosomes are thus essential for the survival of all cellular life forms. Interestingly, ribosomes are made partially from proteins. In a sense, they are also manufacturing themselves. For a long time, all ribosomes of a cell were believed to be the same. Meanwhile, it was discovered that this is not necessarily true. Newer research showed that partially the building blocks of the ribosome can be dynamically changed to regulate the synthesis of specific proteins. In this project, we want to investigate whether a similar dynamic regulation of the ribosomes happens in bacteria. More specifically, we hypothesize that truncated variants of the regular ribosomal proteins are synthesized under certain harsh stress conditions and subsequently integrated into ribosomes. The research on the influence of these adapted ribosomes on the survival of the bacterial cells will thus contribute significantly to our understanding of stress regulation on the level of protein synthesis.

Proteins are the building blocks of all cells. Their production begins with genetic information being transferred from DNA to RNA. Enormous complexes called ribosomes read this information and link amino acids together to form proteins. This makes them essential for the survival of all cellular life forms. Interestingly, since ribosomes themselves are partly composed of proteins, they produce some of their own components. Based on previous data from our research group, in this project, we investigated whether bacteria can form alternative ribosomes under stress. Our hypothesis was that shortened variants of ribosomal proteins arise under severe stress and contribute to survival. One such stress, highly relevant to humans as well, is the elimination of bacteria using antibiotics such as ampicillin, which destroys the cell wall of non-resistant bacteria, causing them to burst. However, there is always a small percentage of bacteria that survive the treatment and contribute to resistance. We call these bacteria persisters. Indeed, we observed that bacterial cells produce a shortened variant of the ribosomal protein L2 (tL2) after treatment with ampicillin. A previous study by a research group in the United States had suggested that this very tL2 could regulate bacterial growth. By applying rigorous controls in our investigations of whether tL2 truly contributes to the survival of persisters, it became apparent that previously published protocols for isolating intact surviving bacteria are insufficient and that proteins from killed cells are co-purified as well. In further experiments, we successfully optimized the existing protocol and were able to clearly demonstrate that ribosomal protein L2 is also released after the destruction of the cell by ampicillin and is processed extracellularly into tL2. Thus, we made an important contribution to research on persistent cells and showed that tL2 is an artifact and cannot contribute to the survival of persistent cells. Although we have not yet been able to confirm our fundamental hypothesis, we subsequently systematically tested the approximately 50 remaining ribosomal proteins using our optimized protocol and identified several interesting candidates that are indeed shortened within persister cells. Further investigations into whether and how these shorter ribosomal proteins contribute to bacterial survival are currently being conducted in our research group. In summary, this research project also exemplifies how scientific work should be carried out. Processes are continuously optimized, results are critically examined, and knowledge is gradually expanded until, through many contributions from different research groups, an increasingly accurate overall picture emerges.