Attachment of the RNA degradosome to the inner cytoplasmic membrane of Escherichia coli prevents wasteful degradation of rRNA intermediates in ribosome assembly.

ABSTRACT: RNase E, an essential endoribonuclease in Escherichia coli, is the key component of the multienzyme RNA degradosome, which is localized to the inner cytoplasmic membrane. Until now, the reason for membrane localization of RNase E was not known. We have analyzed ribosome assembly in the rneΔMTS strain, which expresses a cytoplasmic variant of RNase E (cRNase E) resulting in a cytoplasmic RNA degradosome. In this mutant strain, there is a mild slowdown in the rates of growth and mRNA degradation. Here we document the striking accumulation of intermediates in ribosome assembly in the rneΔMTS strain in which precursors of 16S and 23S rRNA are cleaved by cRNase E. In vitro, we show that ribosomes partially unfolded in low ionic strength buffer are cleaved by RNase E. Mapping of in vivo and in vitro cleavage sites shows that they overlap and that their consensus sequence matches previously mapped RNase E cleavage sites. In vivo, fragments of 16S and 23S rRNA as well as a precursor of 5S rRNA are degraded in a pathway involving polyadenylation and 3’ exonucleases. Since the pathway for rRNA degradation is the same as the pathway for mRNA degradation, the slowdown of mRNA degradation in the rneΔMTS strain could be due to competition by rRNA degradation. Our results strongly suggest that the RNA degradosome participates in the quality control of ribosome assembly and that localization on the inner cytoplasmic membrane protects newly synthesized intermediates from wasteful degradation. Ribosome synthesis is costly. Since growth rate correlates with ribosome synthesis rate, slow growth rate of the rneΔMTS strain is due to the degradation of a proportion of newly synthesized ribosomes. Avoiding wasteful degradation of intermediates in ribosome assembly likely explains why localization of RNase E homologues to the inner cytoplasmic membrane is conserved throughout the γ-Proteobacteria.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Escherichia Coli

TISSUE(S): Cell Culture

SUBMITTER: Carine Froment  

LAB HEAD: Agamemnon J. Carpousis

PROVIDER: PXD038317 | Pride | 2023-01-16

REPOSITORIES: Pride