Periodic Reporting for period 3 - BactRNA (Bacterial small RNAs networks unravelling novel features of transcription and translation)
Reporting period: 2022-09-01 to 2024-02-29
Regulation of gene expression plays a key role in the ability of bacteria to rapidly adapt to changing environments and to colonize extremely diverse habitats. It has been known for decades that multiple regulators, among which two-component systems, control the transcription of many bacterial genes. More recently, the discovery of a plethora of small regulatory RNAs (sRNAs) and the beginning of their characterization highlighted the importance of post-transcriptional regulation in bacterial gene expression. It is now clear that the timely expression of many bacterial genes responds to a complex network of both transcriptional and post-transcriptional regulators. However, the properties of the resulting mixed regulatory circuits on the dynamics of gene expression and in the bacterial adaptive response have been poorly addressed so far.
A first major objective of the BactRNA project is to tackle this question by characterizing the circuits that are formed between two widespread classes of bacterial regulators, the sRNAs and the two-component systems, which act at the post-transcriptional and the transcriptional level, respectively.
In addition, the study of sRNAs also led to major breakthroughs regarding the basic mechanisms of gene expression, and especially the crucial step of translation initiation. For instance, we previously showed that repressor sRNAs can target activating stem-loop structures located within the coding region of mRNAs that promote translation initiation, in striking contrast with the previously recognized inhibitory role of mRNA structures in translation.
We now want to understand how these secondary structures activate translation and how widespread they are among bacterial genes. More generally, a second major objective of this project is to draw an unprecedented map of non-canonical translation initiation events and their regulation by sRNAs.
Overall, we expect this fundamental research project to greatly improve our understanding of how bacteria can so rapidly and successfully adapt to many different environments. On the longer term, results of this project may provide clues towards the development of anti-bacterial strategies.
A first major objective of the BactRNA project is to tackle this question by characterizing the circuits that are formed between two widespread classes of bacterial regulators, the sRNAs and the two-component systems, which act at the post-transcriptional and the transcriptional level, respectively.
In addition, the study of sRNAs also led to major breakthroughs regarding the basic mechanisms of gene expression, and especially the crucial step of translation initiation. For instance, we previously showed that repressor sRNAs can target activating stem-loop structures located within the coding region of mRNAs that promote translation initiation, in striking contrast with the previously recognized inhibitory role of mRNA structures in translation.
We now want to understand how these secondary structures activate translation and how widespread they are among bacterial genes. More generally, a second major objective of this project is to draw an unprecedented map of non-canonical translation initiation events and their regulation by sRNAs.
Overall, we expect this fundamental research project to greatly improve our understanding of how bacteria can so rapidly and successfully adapt to many different environments. On the longer term, results of this project may provide clues towards the development of anti-bacterial strategies.
We have now developed an experimental approach based on reporter fusions with fluorescent proteins allowing us to perform a large-scale screen for small RNAs controlling the synthesis of the regulators of two-component systems. This screen is still in progress, but its first results both confirm known regulations and unravel novel ones, thus identifying mixed regulatory circuits to be investigated in more details as a follow-up.
In addition, we have already started to study several features of specific circuits, among which their role in bacterial virulence or the detailed mechanisms of gene control.
Regarding the regulation of translation initiation by secondary structures, we have first identified cis- and trans-elements involved in the regulation of mRNAs possessing such activating structures. More generally, we also searched for other mRNAs containing this type of structures and found several interesting candidates for which they activate gene expression.
In addition, we have already started to study several features of specific circuits, among which their role in bacterial virulence or the detailed mechanisms of gene control.
Regarding the regulation of translation initiation by secondary structures, we have first identified cis- and trans-elements involved in the regulation of mRNAs possessing such activating structures. More generally, we also searched for other mRNAs containing this type of structures and found several interesting candidates for which they activate gene expression.
Our work provides new illustrations of the sophistication that can be reached in the post-transcriptional control of gene expression in bacteria, such as, for instance, the functional redundancy between two different isoforms of a single sRNA acting on the same target via different mechanisms.
In parallel, we have identified new candidates for control of translation initiation by stem-loop structures located within the coding sequence of mRNAs. Interestingly, these results provide indication that this mode of regulation is conserved in Gram-negative and Gram-positive bacteria.
In the future, we expect that the global approaches that we have launched in this first period will bring important insight into the role of sRNAs in regulatory circuits and into the non-canonical translation initiation events.
In parallel, we have identified new candidates for control of translation initiation by stem-loop structures located within the coding sequence of mRNAs. Interestingly, these results provide indication that this mode of regulation is conserved in Gram-negative and Gram-positive bacteria.
In the future, we expect that the global approaches that we have launched in this first period will bring important insight into the role of sRNAs in regulatory circuits and into the non-canonical translation initiation events.