Biology subjects

Luisi, B.

Publications and source records attributed to Luisi, B..

3 recordsLinked to original sources

bioRxiv2021

Acetylation regulates the oligomerization state and activity of RNase J, the major ribonuclease of Helicobacter pylori

In Helicobacter pylori, post-transcriptional regulation strongly relies on the activity of an RNA degradosome, composed of the essential ribonuclease RNase J and the DEAD-box RNA helicase RhpA. Here, we describe post-translational modifications of this protein complex that affect its activity. Cell-extracted RNase J is acetylated on multiple residues, one of which, K649, strongly impacts RNase J oligomerization, which in turn influences ribonuclease activity. Corroborating the link between oligomerization and activity, mutations targeting K649 and other residues affect the dimerization and in vitro activity of RNase J. Our crystal structure of RNase J reveals three loops that gate access to the active site and rationalizes how oligomerization state influences activity. The acetylated residues of RNase J are important for H. pylori morphology, highlighting that the modifications affect the RNase J cellular function. We propose acetylation as a regulatory level controlling the activity of RNase J and the H. pylori RNA degradosome.

microbiology↗

bioRxiv2020

Antisense oligonucleotides target a nearly invariant structural element from the SARS-CoV-2 genome and drive RNA degradation

RNA structural elements occur in numerous single stranded (+)-sense RNA viruses. The stemloop 2 motif (s2m) is one such element with an unusually high degree of sequence conservation, being found in the 3 UTR in the genomes of many astroviruses, some picornaviruses and noroviruses, and a variety of coronaviruses, including SARS-CoV and SARS-CoV-2. The evolutionary conservation and its occurrence in all viral subgenomic transcripts implicates a key role of s2m in the viral infection cycle. Our findings indicate that the element, while stably folded, can nonetheless be invaded and remodelled spontaneously by antisense oligonucleotides (ASOs) that initiate pairing in exposed loops and trigger efficient sequence-specific RNA cleavage in reporter assays. ASOs also act to inhibit replication in an astrovirus replicon model system in a sequence-specific, dose-dependent manner and inhibit SARS-CoV-2 infection in cell culture. Our results thus permit us to suggest that the s2m element is a site of vulnerability readily targeted by ASOs, which show promise as anti-viral agents.

molecular biology↗

bioRxiv2020

In situ structure of the AcrAB-TolC efflux pump at subnanometer resolution

In Gram-negative bacteria, tripartite efflux pump AcrAB-TolC plays a prominent role in antibiotic resistance. We have used high resolution cryo-ET to visualize the structure of Escherichia coli AcrAB-TolC at a 7 [A] resolution in intact cells. The resulting structures show the detailed architecture of the assembled complex embedded into cell envelope. Interactions with the inner membrane enable crosstalk between AcrB and TolC through AcrA, suggesting that assembly in the native cellular environment is critical for the pump activation mechanism, where the allosteric activating signal is triggered by the alternate binding of AcrA to the lipid membrane and AcrB porter domain. We establish a platform for high resolution in situ structural studies of bacteria efflux pump, which can yield critical information in understanding complex assemblies function. One sentence summaryA 7 [A] in situ structure of the AcrAB-TolC pump from bacteria by cryo-ET reveals mechanisms for active efflux.

biophysics↗