| dc.description.abstracteng | The ability to adapt to changing environmental conditions is essential for every organism to
survive. In bacteria, this adaptation relies on the control of the mRNA synthesis, stability, and
turnover, which allows the expression of different sets of proteins as a response to the external
conditions. In many bacteria, the enzymes responsible for the degradation of the mRNA interact
to form multi-enzyme complexes, the RNA degradosomes. In the Gram-positive model organism
Bacillus subtilis, several binary interactions were detected amongst RNA-related proteins in vivo,
and it has been hypothesized that an RNA-degrading complex exists in this organism. These
interactions involved the endoribonuclease RNase Y, the RNA helicase CshA, the exoribonucleases
RNase J1 and PNPase, and the glycolytic enzymes enolase and phosphofructokinase. Moreover,
the paralogue of RNase J1, RNase J2 was shown to only interact with RNase J1. However, some of
these interactions could not be reproduced and the complex could never be purified as a whole,
questioning the possibility of its existence. In this work, I have studied the subcellular localization
of the components of the RNA degradosome of B. subtilis. The differential localization of each
protein rules out the existence of a stable RNA degradosome. Furthermore, I have studied the
interaction and oligomerization between the paralogous RNases J1 and J2 in vivo, and analyzed
the importance of the C-terminal domain for this interaction. The results confirm the interaction
of the RNases J1 and J2 through the C-terminal domain, and show that they oligomerize as dimers
and tetramers. However, it is possible that the tetramers can only be formed in the presence of
RNA. Moreover, it seems that the RNase J2 cannot interact with RNA on its own.
Although the enzymes of the putative RNA degradosome have been extensively studied, many
questions regarding activity and regulation remain open. Amongst them, the enzyme PNPase is
one of the best studied, since PNPase from Escherichia coli has been studied for many decades. It
has been shown that ATP, c-di-GMP and citrate, amongst others, can regulate its activity.
However, the regulation of the PNPase from B. subtilis is not known. In this work, I have studied
the effect of citrate, c-di-GMP and c-di-AMP, as well as enolase and phosphofructokinase, on the
RNA degradation activity of PNPase. None of these metabolites and enzymes seems to have an
effect on this activity.
Altogether, these findings contribute to a better understanding of the complex picture of RNA
degradation, while opening ways for further investigations. | de |