Biology subjects

Turiak, L.

Publications and source records attributed to Turiak, L..

5 recordsLinked to original sources

bioRxiv2026

Hybrid Tetramers of Wild-Type and Amyloidogenic H88R Transthyretin Challenge the Monomer Paradigm

Hereditary transthyretin amyloidosis (ATTRv) is driven by mutations that destabilize the native tetrameric transthyretin (TTR), promoting monomer formation and amyloid aggregation. The H88R variant has been considered fully monomeric, yet its behavior in heterozygous patients has remained unclear. Here, we demonstrate that H88R TTR can form hybrid tetramers with wild-type chains, both in vitro and in patient sera. We considered all possible tetramers, along with various trimer and dimer constructions, and found that the stability of hybrid tetramers containing one or two mutant chains is not significantly reduced in comparison with of the wild-type tetramer, suggesting a practically unhindered entry for H88R TTR monomers into such hybrid tetrameric assemblies, facilitating their secretion. Mass spectrometry confirms incorporation of H88R chains into tetramers in vitro, and show that the H88R TTR variant is present in the serum of carriers albeit in low concentration. We propose that the scarcity of this variant is the result of its retention in the endoplasmic reticulum and present a model for the association of H88R TTR with the endoplasmic reticule chaperone Binding immunoglobulin Protein (BiP). These findings revise the current monomeric view of H88R TTR, with direct implications for the efficacy of tetramer-stabilizing therapeutics. Our results highlight a delicate balance between cellular retention and hybridization, informing mechanistic understanding and treatment strategies in heterozygous ATTRv patients.

molecular biology↗

bioRxiv2026

Development of a Xylene-Free Sample Preparation Protocol for Quantitative Proteomics of Clinically Relevant Formaldehyde-Fixed Paraffin-Embedded Needle Biopsy Samples

Fresh-frozen tissues are considered the gold standard for proteomic analyses due to superior preservation of protein integrity; however, their use is limited by the logistical and financial requirements of long-term storage. Formaldehyde-fixed paraffin-embedded (FFPE) tissues provide a practical alternative owing to their stability and widespread availability in clinical settings. A critical step in FFPE proteomics is deparaffinization, which traditionally relies on organic solvents such as xylene, along with efficient reversal of formaldehyde-induced crosslinks. In this study, we evaluated multiple FFPE protein extraction and digestion workflows including chaotropic, surfactant-based, and detergent-free approaches in combination with xylene-free deparaffinization strategies, using label-free data-independent acquisition (DIA) LC-MS/MS. Among the tested methods, a chaotropic-, reductant-, and surfactant-free in-solution digestion workflow demonstrated robust protein and peptide recovery. A modified version of this protocol further improved peptide coverage while maintaining comparable protein depth. The applicability of the optimized workflow was assessed using FFPE needle biopsy samples from control, hepatic steatosis, and liver fibrosis groups. Distinct proteomic patterns were observed across conditions, with hepatic steatosis associated with early activation of stress-response pathways, while fibrosis showed evidence suggesting altered lipid metabolism. Overall, this study presents a simple, xylene-free, and MS-compatible workflow for FFPE proteomics that is suitable for low-input clinical samples and may support broader application of archival tissues in proteomic research.

molecular biology↗

bioRxiv2025

A549 tumorigenic and BEAS-2B non-tumorigenic cell line derived small extracellular vesicles show distinct proteomic, N-glycoproteomic and chondroitin/dermatan sulfate profiles

Extracellular vesicles (EVs) are critical mediators of intercellular communication and hold promise as biomarkers and therapeutic targets in cancer, but their molecular alterations remain poorly understood. Protein glycosylation is a frequent post-translational modification; however, most EV studies focus only on proteomics, while mapping glycosylation changes of proteins are still underrepresented. To address this shortcoming, we analyzed the proteomic, N-glycoproteomic, and chondroitin/dermatan sulfate (CS/DS) glycosaminoglycan (GAG) profiles of small EVs (sEVs) derived from A549 lung adenocarcinoma and BEAS-2B non-tumorigenic epithelial cell lines. Principal component analysis and hierarchical clustering revealed that all three profiles are highly dependent on the origin of sEV, highlighting fundamental differences not only at the proteomic but also at the N-glycopeptide and CS/DS levels. Protein expression differences were primarily associated with the upregulation of cell cycle regulation, DNA repair, metabolism, and protein synthesis, while immune-related processes were predominantly downregulated. Proteomics revealed differential expressions of 5 CS proteoglycans, anticipating that their CS profile may also change. N-glycoproteomics highlighted a shift from complex to hybrid N-glycans in cancer sEVs, alongside a significant decrease in fucosylation. Prominent glycoproteins characterized with multiple glycosylation sites included versican, galectin-3-binding protein and laminins. The total amount of CS/DS increased 3.4-fold in cancer sEVs, while the ratio of the two monosulfated disaccharides changed 2-fold, suggesting altered sulfation mechanisms. These findings highlight the potential of N-glycoproteomics and GAG profiling to enhance biomarker discovery and EV-based cancer diagnostics. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=121 SRC="FIGDIR/small/643059v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@c2dfdeorg.highwire.dtl.DTLVardef@1f79452org.highwire.dtl.DTLVardef@12a9dbborg.highwire.dtl.DTLVardef@d992da_HPS_FORMAT_FIGEXP M_FIG C_FIG Proteomic, N-glycoproteomic and chondroitin/dermatan sulfate disaccharide profiles differ between A549 lung adenocarcinoma and BEAS-2B non-tumorigenic epithelial cell derived small extracellular vesicles.

cancer biology↗

bioRxiv2023

ArreSTick Motif is Responsible for GPCR-beta-Arrestin Binding Stability and Extends Phosphorylation-Dependent beta-arrestin Interactions to Non-Receptor Proteins

The binding and function of {beta}-arrestins are regulated by specific phosphorylation motifs present in G protein-coupled receptors (GPCRs). However, the exact arrangement of phosphorylated amino acids responsible for establishing a stable interaction remains unclear. To investigate this pattern, we employed a 1D sequence convolution model trained on a dataset of GPCRs that have established {beta}-arrestin binding properties. This approach allowed us to identify the amino acid pattern required for GPCRs to form stable interactions with {beta}-arrestins. This motif was named "arreSTick." Our data show that the model predicts the strength of the coupling between GPCRs and {beta}-arrestins with high accuracy, as well as the specific location of the interaction within the receptor sequence. Furthermore, we show that the arreSTick pattern is not limited to GPCRs, and is also present in numerous non-receptor proteins. Using a proximity biotinylation assay and mass spectrometry analysis, we demonstrate that the arreSTick motif controls the interaction between numerous non-receptor proteins and {beta}-arrestins. For example, the HIV-1 Tat Specific Factor 1 (HTSF1 or HTATSF1), a nuclear transcription factor, contains the arreSTick pattern, and our data show that its subcellular localization is influenced by its coupling to {beta}-arrestin2. Our findings unveil a broader regulatory role for {beta}-arrestins in phosphorylation-dependent interactions, extending beyond GPCRs to encompass non-receptor proteins as well.

biochemistry↗

bioRxiv2018

Dissection of protein cargo of citrus fruit juice sac cells-derived vesicles reveals heterogeneous transport and extracellular vesicles subpopulations

Cellular vesicles are membrane-enclosed organelles that transport material inside and outside the cell. Plant-derived vesicles are receiving more and more attention due to their potential as nanovectors for the delivery of biologically active substances. We aimed to expand our understanding about the heterogeneity and the protein biocargo of citrus fruit juice sac cell-derived vesicles. Micro- and nano-sized vesicle fractions were isolated from four citrus species, C. sinensis, C. limon, C. paradisi and C. aurantium, characterized using physicochemical methods and protein cargos were compared using label-free quantitative shotgun proteomics. In each sample approximately 600-800 proteins were identified. Orthologues of most of the top-ranking proteins have previously been reported in extracellular vesicles of mammalian origin. High expression of patellin-3-like, clathrin heavy chain, HSPs, 14-3-3 protein, glyceraldehyde-3-phosphate dehydrogenase and fructose-bisphosphate aldolase 6 could be observed in all the vesicle samples while Aquaporin was highly expressed only in nanovesicles. Bioinformatics revealed more than hundred protein orthologues potentially implicated in vesicular trafficking. In particular, the presence of CCV, COPI and COPII coat proteins indicates the presence of highly heterogeneous populations of intracellular transport vesicles. Moreover, the differential expressions of hydrolases and oxidoreductases enabled us to gain insights into their functions in citrus fruit sac cells.

plant biology↗