Biology subjects

Violette, M.

Publications and source records attributed to Violette, M..

5 recordsLinked to original sources

bioRxiv2025

Development and validation of a streamlined workflow for proteomic analysis of proteins and post-translational modifications from dried blood

It is increasingly recognized that the omic analysis of whole blood has applications for precision medicine and disease phenotyping. Despite this realization, whole blood is generally viewed as a challenging analytical matrix in comparison to plasma or serum. Moreover, proteomic analyses of whole blood proteomics have almost exclusively focused on (non)targeted analyses of protein abundances and much less on post-translational modifications (PTMs). Here, we developed a streamlined workflow for processing twenty microliters of venous blood collected by volumetric absorptive microsampling that incorporates serial trypsinization, N-glycopeptide and phosphopeptide enrichment and avoids laborious sample dry-down or cleanup steps. Up to 10,000 analytes (reported as protein groups, glycopeptidoforms and phosphosites) were quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in approximately 2 h of MS acquisition time. Using these methods, we explored the stability of "dried" and "wet" blood proteomes, as well as effects of ex vivo inflammatory stimulus or phosphatase inhibition. Multi-omics factor analysis enabled facile identification of analytes that contributed to inter-individual variability of the blood proteomes, including N-glycopeptides that distinguish immunoglobulin heavy constant alpha 2 allotypes. Collectively, our results help to establish feasibility and best practices for the integrated MS-based quantification of proteins and PTMs from dried blood.

biochemistry↗

bioRxiv2025

Mass spectrometry-based quantification of proteins and post-translational modifications in dried blood: longitudinal sampling of patients with sepsis in Tanzania

The proteomic analysis of blood is routine for disease phenotyping and biomarker development. Whole blood is commonly separated into soluble and cellular fractions. However, this can introduce pre-analytical variability; and analysis of a single component (which is common) may ignore important pathophysiology. We have recently developed methods for the facile processing of dried blood for mass spectrometry-based quantification of the proteome, N-glycoproteome and phosphoproteome. Here, we applied this approach to 38 patients in Tanzania who presented to the hospital with sepsis. Blood was collected on Mitra devices at presentation and 1, 3 and 28-42 days post-enrollment. Processing of 96 total samples was performed in plate-based formats and completed within two days. Approximately 2,000 protein groups and 8,000 post-translational modifications were quantified in 3 LC-MS/MS runs at [~]1.5 hours per sample. Analysis of differential abundance revealed blood proteome signatures of acute phase response and neutrophilic inflammation that partially resolved at the 28-42 day timepoint. Numerous analytes correlated with clinical laboratory values for c-reactive protein and white blood cell counts, as well as the Universal Vital Assessment illness severity score. These datasets serve as proof-of-concept for large scale MS-based (sub)phenotyping of disease using dried blood and are available via the ProteomeXchange consortium (PXD060377).

systems biology↗

bioRxiv2024

Genomic and proteomic characterization of sulfate-reducing symbionts of gutless marine annelids

Sulfate-reducing bacteria (SRB) are widespread in marine and terrestrial environments, where they often form syntrophic associations with bacteria, archaea, and eukaryotes. Among the most intimate of these are multipartite symbioses in gutless marine oligochaete worms, which host SRB and sulfur-oxidizing endosymbionts that engage in a syntrophic exchange of sulfur compounds. Despite decades of research on free-living SRB, the metabolic traits that enable SRB to persist in symbiosis, and how these differ across hosts and environments, remain poorly understood. Here, we show that a globally distributed clade of symbiotic SRB has a conserved core metabolism that diverges markedly from free-living relatives. Using comparative genomics and metaproteomics, we reveal that these endosymbionts retain key traits of SRB such as sulfate reduction, complete oxidation of acetate to CO2, amino acid degradation for nitrogen acquisition, and transport of essential nutrients. However, they exhibit a more oxygen-tolerant metabolism and lack typical nutrient-scavenging mechanisms of free-living SRB. One symbiont-specific trait, the glyoxylate bypass, was consistently expressed in situ and may serve both in reactive oxygen species defence and in biomass generation. The enrichment and expression of oxygen-tolerant pathways, coupled with the loss of nutrient-scavenging functions, indicate specialization to a host-associated, redox-fluctuating environment distinct from that of free-living SRB. Consistent with this shift, symbiont genomes are larger than those of free-living relatives, contrasting with genome reduction in many endosymbionts and reinforcing the importance of metabolic versatility. Our findings provide a framework for understanding how metabolic flexibility enables SRB to persist in long-term multipartite symbioses across diverse marine ecosystems.

microbiology↗

bioRxiv2024

Meta-omics reveals role of photosynthesis in Microbially Induced Carbonate Precipitation at a CO2-rich Geyser

Microbially induced carbonate precipitation (MICP) is a natural process with potential biotechnological applications to address both carbon sequestration and sustainable construction needs. However, our understanding of the microbial processes involved in MICP is limited to a few well-researched pathways such as ureolytic hydrolysis. To expand our knowledge of MICP, we conducted an omics-based study on sedimentary communities from travertine around the CO2-driven Crystal Geyser near Green River, Utah. Using metagenomics and metaproteomics, we identified the community members and potential metabolic pathways involved in MICP. We found variations in microbial community composition between the two sites we sampled, but Rhodobacterales were consistently the most abundant order, including both chemoheterotrophs and anoxygenic phototrophs. We also identified several highly abundant genera of Cyanobacteriales. The dominance of these community members across both sites and the abundant presence of photosynthesis-related proteins suggest that photosynthesis could play a role in MICP at Crystal Geyser. We also found abundant bacterial proteins involved in phosphorous starvation response at both sites suggesting that P-limitation shapes both composition and function of the microbial community driving MICP. Our study emphasizes the possible involvement of photosynthesis in MICP processes at Crystal Geyser and suggests that P-limitation may either hinder or facilitate MICP within the community.

microbiology↗

bioRxiv2023

An intranuclear bacterial parasite of deep-sea musselsexpresses apoptosis inhibitors acquired from its host

Endozoicomonadaceae bacteria are widespread in many marine animals, and generally considered beneficial. Members of one clade, however, Candidatus Endonucleobacter, infect the nuclei of deep-sea mussels, where they replicate to [≥] 80,000 bacteria per nucleus and cause the nuclei to swell to 50 times their original size. How these parasites are able to persist in host nuclei without the cell undergoing apoptosis is not known. We show here that Ca. Endonucleobacter encodes and expresses 7-15 inhibitors of apoptosis (IAPs), proteins previously only known from animals and viruses. Dual RNA-seq transcriptomes of infected nuclei revealed parallel upregulation of Ca. Endonucleobacter IAPs and host caspases, suggesting an arms race between the parasite and host for control of apoptosis. Comparative phylogenetic analyses revealed that Ca. Endonucleobacter acquired IAPs repeatedly through horizontal gene transfer from their hosts in convergent acquisition, possibly mediated by herpes viruses that may infect both the parasite and the host.

microbiology↗