Proteomics最新文献

Bacterial infections have been implicated in shaping the tumor microenvironment (TME), but their effects on cancer cell proteomes remain unexplored. In this study, we analyzed proteomic changes in melanoma (A375) and ovarian cancer (OVCAR3) cell line models following infection with Staphylococcus aureus strain USA300 or Salmonella enterica strain SL1344 using mass spectrometry-based label-free quantitative proteomics. Bacterial infection leads to widespread changes in host protein expression in the cancer cells, with levels of proteins involved in mitochondrial metabolism, RNA processing, and cellular stress response all increasing in relative abundance. In contrast, proteins involved in DNA repair, cytoskeletal structure, vesicle trafficking, and cell cycle regulation were consistently downregulated. The magnitude of the observed changes varied by the cancer cell type. Understanding these interactions may provide new directions for the role of bacteria in tumor progression and therapeutic resistance.

Protein function is dynamically modulated by post-translational modifications (PTMs). Many different types of PTMs can nowadays be identified and quantified at a large scale using mass spectrometry. It is well known that many PTMs have an effect on protein function and cellular processes, and they should be studied not in isolation, but in the holistic context of cellular pathways. This is increasingly facilitated by a wide variety of computational efforts. This review aims to give a systematic overview of tools for pathway-centric analysis of PTM data and critically evaluate the state of play in this research field. Starting from databases that make up the foundational prior knowledge, we follow typical steps that an analytical workflow might contain, including pathway enrichment analysis, algorithms for pathway reconstruction, and the integration and visualization of results. We then reflect on common limitations of all existing tools and give our opinion on future directions that we think are currently most desirable.

Mass spectrometry (MS) offers robust, label-free approaches for characterizing ligand–protein interactions through two main strategies: affinity-based and stability-based assays. However, their application to membrane proteins (MPs)—a major class of drug targets—has been limited by challenges such as structural complexity, low native expression, incomplete trypsin digestion, and poor compatibility with detergent-based MS protocols. Recent progress has advanced the field along two complementary fronts. First, innovations in MS methodology, including native MS, nativeomics, solution-phase thermochemistry, and ion mobility-mass spectrometry (IM-MS), have improved the ability to preserve intact assemblies, capture co-bound lipids and ligands, and resolve conformational and energetic landscapes of MPs. Second, advances in MP solubilization and stabilization, through tailored detergent architectures, MS-compatible detergents, and membrane mimetic (MM) systems—such as nanodiscs, peptidiscs, and styrene–maleic acid (SMA) polymers—have created more native-like environments that maintain functional conformations and ligand-binding sites, enabling integration of MPs into high-throughput MS platforms for ligand screening. This review outlines key affinity- and stability-based MS approaches for MPs and highlights how advances in MS methodology and solubilization strategies are extending their scope, positioning MS and MM as an increasingly powerful platform for high-throughput discovery of MP–ligand interactions.

Lysosomes constitute the main degradative organelle of most eukaryotic cells and are capable of breaking down a wide spectrum of biomolecules, including proteins, lipids, glycans, and DNA/RNA. They play crucial roles in the regulation of cellular homeostasis, acting as metabolic signaling centers for the correlation of nutrient availability and biosynthetic processes. The lysosome's importance is highlighted by several human diseases associated with its dysfunction, including both early- and late-onset conditions, dependent on the level of functional impairment. Lysosomal biogenesis presents a multi-step process consisting of various delivery routes for its individual constituents, enabling strict activity control of the currently known ∼60 lysosomal hydrolases to prevent cellular self-digestion and proper assembly of the lysosomal membrane. In this review, we recapitulate the contribution of mass spectrometry (MS)-based proteomics to the characterization of lysosomal biogenesis in the last two decades. The enrichment and proteomic analysis of lysosomes and lysosomal proteins played an invaluable role for the investigation of lysosomes, encompassing the control of lysosomal gene expression, the characterization of sorting/trafficking processes, and the assignment of lysosomal proteins. This has resulted so far in the definition of ∼350 proteins which have been identified to be located in/at lysosomes or are of crucial importance for their function.

Extracellular vesicles (EVs) and particles (EPs) are diverse micro- and nanoparticles that circulate in bodily fluids and can attach to, or be deposited onto, the extracellular matrix (ECM) and other surfaces. To date, the nomenclature and classification of matrix-bound or matrix-associated EVs and EPs (MEVPs) have been unclear, largely due to a lack of consensus guidelines and a relatively miniscule amount of received attention in comparison to EVs found in fluids. Recently, there has been a growing appreciation for several subtypes of MEVPs and their roles in applications ranging from wound healing to metastasis. However, progress in these fields has largely been achieved in silos, with minimal consideration for overlap or complementary function between different MEVPs. In this article, we briefly describe this growing field with a focus on several MEVP subtypes and the lack of consensus, then discuss challenges and opportunities in improving MEVP isolation and characterization. Importantly, proteomic analyses of these unique MEVPs will be crucial in promoting rigor, reproducibility, and understanding in this exciting new field.

Although oxidative stress is a well-established driver of neurodegeneration, it remains poorly understood as to how the global cysteine (Cys) proteome is remodeled under oxidative stress conditions. Proteins with aberrantly modified cysteines in response to oxidative stress can induce and exacerbate neurodegeneration, contributing to disorders like Alzheimer's, Parkinson's, frontotemporal dementia, and amyotrophic lateral sclerosis. In this study, we induced oxidative stress in SH-SY5Y neuronal cells by subjecting them to the neurotoxin 6-hydroxydopamine (6-OHDA). To identify proteins with altered cysteine oxidation or PTM status, we used a desthiobiotin iodoacetamide (DBIA) probe, which selectively labels cysteines with unmodified and preserved thiols. Using these unbiased chemoproteomic strategies, we identified proteins with reduced Cys reactivity to DBIA in response to 6-OHDA-induced oxidative stress. Many of these proteins are critically involved in biological processes linked to cell stress responses (e.g., mitochondrial oxidative stress and apoptosis). Furthermore, we found that two key Cys on UCHL1 (a deubiquitinase critically involved in neurodegeneration) exhibited enhanced reactivity under oxidative stress conditions. Our study defines the remodeling of the Cys proteome under 6-OHDA-induced oxidative stress conditions. Furthermore, these findings suggest potential cysteine-mediated regulatory mechanisms in response to oxidative stress, providing a valuable resource for further exploration of cysteine modifications in the context of neurodegenerative signaling.