Biology subjects

Guarneros, G.

Publications and source records attributed to Guarneros, G..

4 recordsLinked to original sources

bioRxiv2025

Halves and fragments derived of tRNAs in Escherichia coli are selectively associated with 30S ribosomal subunits and in the cytosol

The generation of tRNA halves and fragments (tsRNAs) has been associated with stressful growth conditions in eukaryotes, but reports on tsRNAs in bacteria are scarce. Here, we demonstrate that the presence of tsRNAs in Escherichia coli depends on active translation, and that they are found in the 30S ribosomal subunit and the ribosomal-free fraction, but not in the 50S subunits, 70S ribosomes or polysome fractions. However, upon dissociation into subunits at a low magnesium concentration, some of the tRNAs present in the monosomal and polysomal fractions are processed into tsRNAs. RNA-seq analyses tsRNA fractions revealed that all tRNA species in the cell were processed into fragment profiles that varied widely for each tRNA. The E. coli CP78 strain contains an unusually high concentration of tsRNAAsnGUU, but it is likely that only a fraction of this participates in translation. These tRNAs, along with others in the cell, were released from the ribosomes and processed into tsRNAs. The tRNAs in the ribosomal-free fraction appear to be cleaved by the same RNase that is active in ribosomes. We propose that tsRNAs are generated as an initial decay step for tRNAs remaining on ribosomes following translation arrest. However, tsRNAs may also have other functions.

molecular biology↗

bioRxiv2022

Transcriptional analysis in bacteriophage Fc02 of Pseudomonas aeruginosa revealed two overlapping genes with exclusion activity

Little is known about the gene expression program during transition from lysogenic to lytic cycles of temperate bacteriophages in Pseudomonas aeruginosa. To investigate this issue, we developed a thermo-sensitive repressor mutant in a lysogen and analyzed the phage transcriptional program by strand-specific RNA-Seq before and after thermo-induction. As expected, the repressor gene located on the phage DNA forward strand, is transcribed in the lysogen at the permissive temperature of 30{degrees}C. Upstream the promoter gene, we noticed the presence of two overlapped ORFs apparently in the same transcript, one ORF is a gene that encodes a protein of 7.9 kDa mediating exclusion of various super-infecting phages. The other ORF, placed in an alternate reading frame, with a possible AUG initiation codon at 25 n downstream the AUG of the first gene, is expected to encode a 20.7 kDa polypeptide of yet unknown function. Upon lifting repression at 40{degrees}C, starts transcription of an operon, involved in the lytic cycle from a promoter on the reverse phage DNA strand. The first gene in the operon is a homolog of the antirepresor ner, a common gene in the lysis-lysogeny regulation region of other phages. Interestingly, the next gene after ner is gene10 that on the reverse strand, overlaps the overlapped gene olg1 on the forward strand. Curiously, gene 10 expression also shows superinfection exclusion. Strand-specific RNA-Seq also has uncover the transcription succession of gene modules expressed during the phage lytic stage.

microbiology↗

bioRxiv2021

Molecular evolution of the anti-CRISPR family AcrIF7 reveals conservation despite mutation tolerance

Anti-CRISPRs are proteins capable of blocking CRISPR-Cas systems and typically encoded in mobile genetic elements. Since their discovery, numerous anti-CRISPR families have been identified. However, little is known about the distribution and sequence diversity of members within a family, nor how these traits influence the anti-CRISPRs function and evolution. Here we use AcrIF7 to explore the dissemination and molecular evolution of an anti-CRISPR family. We uncovered five sub-clusters and prevalent anti-CRISPR variants within the group. Remarkably, AcrIF7 homologs display high similarity despite their broad geographical, ecological and temporal distribution. Although mainly associated with Pseudomonas aeruginosa, AcrIF7 was identified in distinct genetic backgrounds indicating horizontal dissemination, primarily by phages. Using mutagenesis, we recreated variation observed in databases but also extended the sequence diversity of the group. Characterisation of the variants identified residues key for the anti-CRISPR function and other contributing to its mutational tolerance. Moreover, molecular docking revealed that variants with affected function lose key interactions with its CRISPR-Cas target. Analysis of publicly available data and the generated variants suggests that the dominant AcrIF7 variant corresponds to the minimal and optimal anti-CRISPR selected in the family. Our study provides a blueprint to investigate the molecular evolution of anti-CRISPR families.

microbiology↗

bioRxiv2020

Dynamics of infection in a novel group of promiscuous phages and hosts of multiple bacterial genera retrieved from river communities

Phages are generally described as species- or even strain-specific viruses, implying an inherent limitation for some to be maintained and spread in diverse bacterial communities. Moreover, phage isolation and host range determination rarely consider the phage ecological context, likely biasing our notion on phage specificity. Here we identified and characterized a novel group of promiscuous phages existing in rivers by using diverse bacteria isolated from the same samples, and then used this biological system to investigate infection dynamics in distantly related hosts. We assembled a diverse collection of over 600 native bacterial strains and used them to isolate six podophages, named Atoyac, from different geographic origin and capable of infecting six genera in the Gammaproteobacteria. Atoyac phage genomes are highly similar to each other but not to those currently available in the genome and metagenome public databases. Detailed comparison of the phages infectivity in diverse hosts and trough hundreds of interactions revealed variation in plating efficiency amongst bacterial genera, implying a cost associated with infection of distant hosts, and between phages, despite their sequence similarity. We show, through experimental evolution in single or alternate hosts of different genera, that plaque production efficiency is highly dynamic and tends towards optimization in hosts rendering low plaque formation. Complex adaptation outcomes observed in the evolution experiments differed between highly similar phages and suggest that propagation in multiple hosts may be key to maintain promiscuity in some viruses. Our study expands our knowledge of the virosphere and uncovers bacteria-phage interactions overlooked in natural systems. ImportanceIn natural environments, phages co-exist and interact with a broad variety of bacteria, posing a conundrum for narrow-host-range phages maintenance in diverse communities. This context is rarely considered in the study of host-phage interactions, typically focused on narrow-host-range viruses and their infectivity in target bacteria isolated from sources distinct to where the phages were retrieved from. By studying phage-host interactions in bacteria and viruses isolated from river microbial communities, we show that novel phages with promiscuous host range encompassing multiple bacterial genera can be found in the environment. Assessment of hundreds of interactions in diverse hosts revealed that similar phages exhibit different infection efficiency and adaptation patterns. Understanding host range is fundamental in our knowledge of bacteria-phage interactions and their impact in microbial communities. The dynamic nature of phage promiscuity revealed in our study has implications in different aspects of phage research such as horizontal gene transfer or phage therapy.

microbiology↗