Biology subjects

Meyer, M. M.

Publications and source records attributed to Meyer, M. M..

11 recordsLinked to original sources

bioRxiv2026

Translation Inhibiting Antibiotics Induce a Strain Specific asRNA Regulating Purine Metabolism in S. pneumoniae TIGR4

Translation inhibiting antibiotics (TIA) are an important class of antibiotics that elicit a wide variety of transcriptional responses. In this work we characterize the transcriptional responses of Streptococcus pneumoniae TIGR4, via RNA-seq, 5-, and 3-end mapping, to sub-MIC levels of three TIA, chloramphenicol, tetracycline, and kasugamycin. We find that kasugamycin (initiation inhibitor) displays a distinct transcriptomic profile compared to chloramphenicol and tetracycline (elongation inhibitors). However, genes in nucleotide metabolism were consistently downregulated. We also detected a TIA induced antisense transcript complementary to downregulated genes for purine metabolism, which we term SP_0835as. Mutating the promoter for SP_0835as eliminates both induction of SP_0835as and repression of the complementary genes in response to chloramphenicol. The mutated strain also displays slower growth rates than control strains in conditions that strongly induce SP_0835as, implying biologically relevant activity. Furthermore, the SP_0835as promoter straddles the boundary between SP_0835 and an insertion sequence (ISL3) immediately downstream. A survey of S. pneumoniae genomes shows the promoter is present in several phylogenetically distinct strain clusters. However, other common laboratory strains of S. pneumoniae lack the promoter, SP_0835as expression, and TIA repression of the complementary genes. Thus, SP_0835as represents a nascent RNA regulator that significantly remodels the metabolic response to TIA.

microbiology↗

bioRxiv2026

Electrical Readout Strategies of GFET Biosensors for Real-World Requirements

Graphene Field-Effect Transistors (GFETs) are increasingly employed as biochemical sensors due to their exceptional electronic properties, surface sensitivity, and potential for miniaturization. A critical challenge in deploying GFETs is determining the optimal electrical readout strategy. GFETs are typically operated with either of two modalities: one measuring current in real time (amperometric) and the other monitoring the change in voltage for charge neutrality (potential potentiometric). Here, we undertake a systematic study of the two modalities to determine their relative advantages/disadvantages towards guiding the future use of GFETs in sensing. We focus on viral proteins in wastewater, given the matrixs complexity and the growing interest in the field of wastewater surveillance. Our results show that transconductance offers far superior limits of detection (LOD) but suffers from limited reproducibility, a narrower dynamic range, and is ineffective for some viral proteins. In comparison, we find that Dirac point tracking offers higher reproducibility and superior robustness, but at a higher LOD. Interestingly, both techniques exhibit similar sensitivity, highlighting the importance of the aptamers employed. Systematic experiments also help explain differences in dynamic range and limited functionality in detecting some proteins, resulting from hidden electrophoresis, and shifting the high transconductance point away from the active region. Thus, our findings provide crucial considerations for designing and operating resilient graphene biosensors suitable for real-time pathogen monitoring in environmental scenarios.

biophysics↗

bioRxiv2025

Analysis of 3'-seq data from multiple E. coli studies identifies diverging results sets and raw data characteristics despite similar collection conditions

3-prime end sequencing (3-seq) is a high-throughput sequencing technique that is used to specifically quantify the changes in 3-end formation of transcripts in bacterial cells, which is increasingly being utilized to address fundamental questions regarding transcription termination and pausing across a range of different bacterial species. However, the growing number of 3-seq studies is accompanied by an increase in study-specific 3-seq data analysis approaches. Thus, differences in a number of factors including: experimental design, data collection approaches, analysis methodologies, and interpretation decisions, make it challenging to confidently compare results derived from different studies, even those that were performed on the same organism. To assess the potential severity of these discrepancies, we used PIPETS, a statistically robust and genome-annotation agnostic 3-seq analysis package, to study Escherichia coli 3-seq data sets from three different groups collected under similar conditions. By using a consistent analysis and results interpretation approach, we identified large disparities in the characteristics of the raw 3-seq data between each of the studies, despite all three studies using the same strain and very similar reported experimental conditions. Additionally, we found strand-specific inconsistencies, with some data sets having reference strand 3-seq read coverage distributions that differed greatly from the complement strand within the same replicate. Finally, when the 3-seq distribution profiles of the three E. coli studies are compared to studies from four additional bacteria, we identified 3-seq results clustering patterns that are not explained by phylogenetic similarity between organisms. With the large differences seen between data sets from the same organism as well as the inconsistencies seen between replicates from the same data sets, we urge the field to reconsider the assumptions around 3-seq data homogeneity and move towards consistent analysis approaches, and cautious interpretation of the data. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=79 SRC="FIGDIR/small/658996v1_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@ca8c67org.highwire.dtl.DTLVardef@1c7c7d4org.highwire.dtl.DTLVardef@11047cdorg.highwire.dtl.DTLVardef@1da1791_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2025

Fitness landscapes for an FMN riboswitch in S. pneumoniae reveals complex interplay between antibiotic inhibitors and the native ligand as well as insights into the mechanism of action

RNA-based control of gene expression is common across all domains of life, yet shows extraordinary variety in terms of environmental cues recognized, mechanisms of action, required protein factors, and magnitude of gene expression changes. The fitness benefit conferred by RNA-mediated regulation is often subtle and challenging to measure and may be highly influenced by the environment. Yet selective forces are sufficient to generate and maintain complex RNA structures that allow rapid and robust response to stimuli. In this work we perform deep mutational scanning to measure the relative fitness conferred by all single mutants of an FMN riboswitch to the opportunistic pathogen Streptococcus pneumoniae under seven different conditions. We find that in standard culture conditions the RNA sequence is not under selective pressure, however the dynamic conditions encountered during mouse lung infection reveal an sequence that is under weak purifying selection, but robust to strongly deleterious mutations. Furthermore, assessment of fitness in the presence of two different targeting antibiotics exposes diverging adaptive landscapes that reveal suites of mutations conferring either constitutive activation or repression of gene expression. Analysis of these landscapes highlights a complex relationship between antibiotic concentration and the fitness benefit conferred by individual mutations as well as illuminates the mechanism of action for this RNA with lessons broadly applicable to interpretation of bacterial transcriptomics.

genetics↗

bioRxiv2024

Assessing the conservation and targets of putative sRNAs in Streptococcus pneumoniae

RNA regulators are often found in regulatory networks and mediate growth and virulence in bacteria. Small RNAs (sRNAs) are non-coding RNAs that modulate translation initiation and mRNA degradation by base-pairing. To better understand the role of sRNAs in pathogenicity several studies identified sRNAs in Streptococcus pneumoniae; however, little functional characterization has followed. This studys goals are: 1) survey putative sRNAs in S. pneumoniae; 2) assess the conservation of these sRNAs; and 3) examine their predicted targets. Three previous studies in S. pneumoniae identified 287 putative sRNAs by high-throughput sequencing. This study narrows the candidates to a list of 58 putative sRNAs. BLAST analysis indicates that the 58 sequences are highly conserved across the S. pneumoniae pangenome, and 25 of them are identified sporadically in other Streptococcus species. However, only 2 have corresponding sequences identified across several Streptococcus species. We used four RNA-target prediction programs to predict targets for each of the 58 putative sRNAs. Across all probable predictions, six sRNAs have overlapping targets predicted by multiple programs, four of which target numerous transposase encoding transcripts. sRNAs targeting transposase genes display nearly identical and perfect base-pairing. One sRNA, M63 (Spd_sr37), has several probable targets in the CcpA regulon, a network responsible for global catabolite repression, suggesting a possible biological function in carbon metabolism control. Each M63-target interaction exhibits unique base-pairing increasing confidence in the biological relevance of the result. This study produces a list of S. pneumoniae putative sRNAs whose predicted targets suggest functional significance in transposon and carbon metabolism regulation. ImportancePrevious studies identified many small RNA candidates in Streptococcus pneumoniae, several of which were hypothesized to play a role in S. pneumoniae virulence. Due to the differing sequencing methods, diverse inclusion criteria, S. pneumoniae strain differences, as well as limited follow-up since, it is unclear to what extent candidates identified in different studies have overlapping sequences and functions, and their biological relevance remains ambiguous. This research aims to consolidate the candidate sRNAs across these studies and focuses attention on those that are likely to be regulatory and associated with virulence. This studys findings enhance our knowledge of the conservation of small regulatory RNAs across the many Streptococcus pneumoniae strains and highlights a handful that appear likely to have a role in growth or virulence.

bioinformatics↗

bioRxiv2024

PIPETS: A statistically robust, gene-annotation agnostic analysis method to study bacterial termination using 3'-end sequencing.

BackgroundOver the last decade the drop in short-read sequencing costs has allowed experimental techniques utilizing sequencing to address specific biological questions to proliferate, oftentimes outpacing standardized or effiective analysis approaches for the data generated. There are growing amounts of bacterial 3-end sequencing data, yet there is currently no commonly accepted analysis methodology for this datatype. Most data analysis approaches are somewhat ad hoc and, despite the presence of substantial signal within annotated genes, focus on genomic regions outside the annotated genes (e.g. 3 or 5 UTRs). Furthermore, the lack of consistent systematic analysis approaches, as well as the absence of genome-wide ground truth data, make it impossible to compare conclusions generated by diffierent labs, using diffierent organisms. ResultsWe present PIPETS, (Poisson Identification of PEaks from Term-Seq data), an R package available on Bioconductor that provides a novel analysis method for 3-end sequencing data. PIPETS is a statistically informed, gene-annotation agnostic methodology. Across two diffierent datasets from two diffierent organisms, PIPETS identified significant 3-end termination signal across a wider range of annotated genomic contexts than existing analysis approaches, suggesting that existing approaches may miss biologically relevant signal. Furthermore, assessment of the previously called 3-end positions not captured by PIPETS showed that they were uniformly very low coverage. ConclusionsPIPETS provides a broadly applicable platform to explore and analyze 3-end sequencing data sets from across diffierent organisms. It requires only the 3-end sequencing data, and is broadly accessible to non-expert users.

bioinformatics↗

bioRxiv2023

Evaluation of the fitness benefit conferred by RNA cis-regulators to Streptococcus pneumoniae during infection

Bacteria have evolved complex transcriptional regulatory networks, as well as many diverse regulatory strategies at the RNA level, to enable more efficient use of metabolic resources and rapid response to changing conditions. However, most RNA-based regulatory mechanisms are not well conserved across different bacterial species despite controlling genes important for virulence or essential biosynthetic processes. Here, we characterize the activity of, and assess the fitness benefit conferred by, twelve cis-acting regulatory RNAs (including several riboswitches and a T-box), in the opportunistic pathogen Streptococcus pneumoniae TIGR4. By evaluating native locus mutants of each regulator that result in constitutively active and repressed expression, we establish that growth defects in planktonic culture are associated with constitutive repression of gene expression, while constitutive activation of gene expression is rarely deleterious. In contrast, in mouse nasal carriage and pneumonia models, strains with both constitutively active and repressed gene expression are significantly less fit than matched control strains. Furthermore, two RNA-regulated pathways, FMN synthesis/transport and pyrimidine synthesis/transport display the greatest sensitivity to mis-regulation or constitutive gene repression in both planktonic culture and in vivo environments. Our findings suggest that despite lack of an obvious phenotypes associated with gene over-expression in vitro, the fitness benefit conferred on bacteria via fine-tuned metabolic regulation through cis-acting regulatory RNAs is often substantial in vivo, and therefore easily sufficient to drive the evolution and maintenance of diverse RNA regulatory mechanisms.

microbiology↗

bioRxiv2019

Comparative metatranscriptomics of periodontitis supports a common polymicrobial shift in metabolic function and identifies novel putative disease-associated ncRNAs

Periodontitis is an inflammatory disease that deteriorates bone supporting teeth afflicting [~]743 million people worldwide. Bacterial communities associated with disease have been classified into red, orange, purple, blue, green, and yellow complexes based on their roles in the periodontal pocket. Previous metagenomic and metatranscriptomics analyses suggest a common shift in metabolic signatures in disease vs. healthy communities with up-regulated processes including pyruvate fermentation, histidine degradation, amino acid metabolism, TonB-dependent receptors. In this work, we examine existing metatranscriptome datasets to identify the commonly differentially expressed transcripts and potential underlying RNA regulatory mechanisms behind the metabolic shifts. Raw RNA-seq reads from three studies (including 49 healthy and 48 periodontitis samples) were assembled into transcripts de novo. Analyses revealed 859 differentially expressed (DE) transcripts, 675 more- and 174 less-expressed. Only [~]20% of the DE transcripts originate from the pathogenic red/orange complexes, and [~]50% originate from organisms unaffiliated with a complex. Comparison of expression profiles revealed variations among disease samples; while specific metabolic processes are commonly up-regulated, the underlying organisms are diverse both within and across disease associated communities. Surveying DE transcripts for known ncRNAs from the Rfam database identified a large number of tRNAs and tmRNAs as well as riboswitches (FMN, glycine, lysine, and SAM) in more prevalent transcripts and the cobalamin riboswitch in both more and less prevalent transcripts. In silico discovery identified many putative ncRNAs in DE transcripts. We report 15 such putative ncRNAs having promising covariation in the predicted secondary structure and interesting genomic context. Seven of these are antisense of ribosomal proteins that are novel and may involve maintaining ribosomal protein stoichiometry during the disease associated metabolic shift. Our findings describe the role of organisms previously unaffiliated with disease and identify the commonality in progression of disease across three metatranscriptomic studies. We find that although the communities are diverse between individuals, the switch in metabolic signatures characteristic of disease is typically achieved through the contributions of several community members. Furthermore, we identify many ncRNAs (both known and putative) which may facilitate the metabolic shifts associated with periodontitis.

microbiology↗

bioRxiv2019

Regulatory context drives conservation of glycine riboswitch aptamers

In comparison to protein coding sequences, the impact of mutation and natural selection on the sequence and function of non-coding (ncRNA) genes is not well understood. Many ncRNA genes are narrowly distributed to only a few organisms, and appear to be rapidly evolving. Compared to protein coding sequences, there are many challenges associated with assessment of ncRNAs that are not well addressed by conventional phylogenetic approaches, including: short sequence length, lack of primary sequence conservation, and the importance of secondary structure for biological function. Riboswitches are structured ncRNAs that directly interact with small molecules to regulate gene expression in bacteria. They typically consist of a ligand-binding domain (aptamer) whose folding changes drive changes in gene expression. The glycine riboswitch is among the most well-studied due to the widespread occurrence of a tandem aptamer arrangement (tandem), wherein two homologous aptamers interact with glycine and each other to regulate gene expression. However, a significant proportion of glycine riboswitches are comprised of single aptamers (singleton). Here we use graph clustering to circumvent the limitations of traditional phylogenetic analysis when studying the relationship between the tandem and singleton glycine aptamers. Graph clustering enables a broader range of pairwise comparison measures to be used to assess aptamer similarity. Using this approach, we show that one aptamer of the tandem glycine riboswitch pair is typically much more highly conserved, and that which aptamer is conserved depends on the regulated gene. Furthermore, our analysis also reveals that singleton aptamers are more similar to either the first or second tandem aptamer, again based on the regulated gene. Taken together, our findings suggest that tandem glycine riboswitches degrade into functional singletons, with the regulated gene(s) dictating which glycine-binding aptamer is conserved.\n\nAuthor SummaryThe glycine riboswitch is a ncRNA responsible for the regulation of several distinct gene sets in bacteria that is found with either one (singleton) or two (tandem) aptamers, each of which directly senses glycine. Which aptamer is more important for gene-regulation, and the functional difference between tandem and singleton aptamers, are long-standing questions in the riboswitch field. Like many biologically functional RNAs, glycine aptamers require a specific 3D folded conformation. Thus, they have low primary sequence similarity across distantly related homologs, and large changes in sequence length that make creation and analysis of accurate multiple sequence alignments challenging. To better understand the relationship between tandem and singleton aptamers, we used a graph clustering approach that allows us to compare the similarity of aptamers using metrics that measure both sequence and structure similarity. Our investigation reveals that in tandem glycine riboswitches, one aptamer is more highly conserved than the other, and which aptamer is conserved depends on what gene(s) are regulated. Moreover, we find that many singleton glycine riboswitches likely originate from tandem riboswitches in which the ligand-binding site of the non-conserved aptamer has degraded over time.

bioinformatics↗

bioRxiv2018

Changes in endosymbiont complexity drive host-level compensatory adaptations in cicadas

For insects that depend on one or more bacterial endosymbionts for survival, it is critical that these bacteria are faithfully transmitted between insect generations. Cicadas harbor two essential bacterial endosymbionts, Sulcia muelleri and Hodgkinia cicadicola. In some cicada species, Hodgkinia has fragmented into multiple distinct cellular and genomic lineages that can differ in abundance by more than two orders of magnitude. This complexity presents a potential problem for the host cicada, because low-abundance-but-essential Hodgkinia lineages risk being lost during the symbiont transmission bottleneck from mother to egg. Here we show that all cicada eggs seem to receive the full complement of Hodgkinia lineages, and that in cicadas with more complex Hodgkinia this outcome is achieved by increasing the number of Hodgkinia cells transmitted by up to six-fold. We further show that cicada species with varying Hodgkinia complexity do not visibly alter their transmission mechanism at the resolution of cell biological structures. Together these data suggest that a major cicada adaptation to changes in endosymbiont complexity is an increase in the number of Hodgkinia cells transmitted to each egg. We hypothesize that the requirement to increase the symbiont titer is one of the costs associated with Hodgkinia fragmentation.

evolutionary biology↗

bioRxiv2018

The Transcriptional landscape of Streptococcus pneumoniae reveals a complex operon architecture and abundant riboregulation critical for growth and virulence

Efficient and highly organized transcription initiation and termination is fundamental to an organisms ability to survive, proliferate, and quickly respond to its environment. Over the last decade, our simplistic outlook of bacterial transcriptional regulation and architecture has evolved to include stimulus-responsive regulation by untranslated RNA and the formation of alternate transcriptional units. In this study, we map the transcriptional landscape of the bacterial pathogen Streptococcus pneumoniae by applying a combination of high-throughput RNA-sequencing techniques. Our study reveals a complex transcriptome wherein environment-respondent alternate transcriptional units are observed within operons stemming from internal transcription start sites (TSS) and transcription terminators (TTS) suggesting that more fine-tuning of regulation occurs than previously thought. Additionally, we identify many putative cis-regulatory RNA elements and riboswitches within 5-untranslated regions (5-UTR) of genes. By integrating TSSs and TTSs with independently collected RNA-Seq datasets from a variety of conditions, we establish the response of these regulators to changes in growth conditions and validate several of them. Furthermore, to determine the importance of ribo-regulation by 5-UTR elements for in vivo virulence, we show that the pyrR regulatory element is essential for survival, successful colonization and infection in mice suggesting that such RNA elements are potential drug targets. Importantly, we show that our approach of combining high-throughput sequencing with in vivo experiments can reconstruct a global understanding of regulation, but also pave the way for discovery of compounds that target (ribo-) regulators to mitigate virulence and antibiotic resistance.

microbiology↗