Periodic Reporting for period 1 - OMRP (O-Glc-NAc modifications of RNA-binding proteins)
Période du rapport: 2023-07-01 au 2025-08-31
Proteins are central to virtually every biological process, and their activity is often fine-tuned by chemical modifications after synthesis. One such regulatory mechanism is O-GlcNAcylation, the attachment of a single N-acetylglucosamine sugar molecule (O-GlcNAc) to specific amino acid residues. This dynamic and reversible modification is catalyzed by two key enzymes: O-GlcNAc transferase (OGT), which adds the modification, and O-GlcNAcase (OGA), which removes it.
Recent studies indicate that a substantial proportion of RNA-binding proteins—key regulators of RNA processing, stability, and translation—undergo O-GlcNAcylation. Notably, the RNA-binding protein TDP-43, when O-GlcNAcylated, shows reduced aggregation propensity. Aggregation of TDP-43 into insoluble inclusions is a hallmark of several neurodegenerative conditions, including amyotrophic lateral sclerosis (ALS) and certain dementias, and contributes to neuronal dysfunction and loss.
Pharmacological inhibition of OGA has emerged as an effective and safe strategy to increase cellular O-GlcNAcylation, with promising results in preventing pathological protein aggregation in cellular and animal models. However, the broader effects of elevated O-GlcNAcylation on the structure, function, and interaction networks of RNA-binding proteins remain poorly understood.
The OMRP project will systematically investigate how enhanced O-GlcNAcylation alters the biochemical and functional properties of RNA-binding proteins, with a particular focus on those implicated in neurodegeneration. By combining advanced proteomics, biochemical assays, and functional analyses, this research will generate fundamental insights into the role of O-GlcNAcylation in protein homeostasis.
Recent studies indicate that a substantial proportion of RNA-binding proteins—key regulators of RNA processing, stability, and translation—undergo O-GlcNAcylation. Notably, the RNA-binding protein TDP-43, when O-GlcNAcylated, shows reduced aggregation propensity. Aggregation of TDP-43 into insoluble inclusions is a hallmark of several neurodegenerative conditions, including amyotrophic lateral sclerosis (ALS) and certain dementias, and contributes to neuronal dysfunction and loss.
Pharmacological inhibition of OGA has emerged as an effective and safe strategy to increase cellular O-GlcNAcylation, with promising results in preventing pathological protein aggregation in cellular and animal models. However, the broader effects of elevated O-GlcNAcylation on the structure, function, and interaction networks of RNA-binding proteins remain poorly understood.
The OMRP project will systematically investigate how enhanced O-GlcNAcylation alters the biochemical and functional properties of RNA-binding proteins, with a particular focus on those implicated in neurodegeneration. By combining advanced proteomics, biochemical assays, and functional analyses, this research will generate fundamental insights into the role of O-GlcNAcylation in protein homeostasis.
We conducted an integrated multi-omics approach combining proteomics and transcriptomics to investigate cellular changes associated with elevated O-GlcNAcylation. Using advanced mass spectrometry–based proteome profiling and RNA sequencing, we systematically identified pathways and molecular networks affected by increased protein O-GlcNAc modification. This analysis revealed specific alterations in RNA-binding proteins, stress response factors, and signaling regulators. To ensure robustness, key findings were validated through complementary experimental techniques, including immunofluorescent staining for cellular localization and quantitative ARTR-seq to confirm transcript-level changes. Together, these efforts provided a comprehensive view of how O-GlcNAcylation remodels cellular physiology and identified candidate proteins and pathways that may play a critical role in disease-relevant processes.
The results of this study provide new mechanistic insights into how O-GlcNAcase inhibitors influence cellular pathways and how elevated glucose levels reshape protein regulation through O-GlcNAcylation. By integrating proteomic and transcriptomic analyses, we uncovered key molecular changes that link altered O-GlcNAc cycling to cellular stress responses, RNA metabolism, and protein aggregation. These findings not only refine our understanding of the mode of action of O-GlcNAcase inhibitors but also highlight broader metabolic adaptations triggered by high-glucose environments.
Potential impacts:
Advancement of therapeutic strategies: The results can inform the rational design and optimization of O-GlcNAcase inhibitors as potential therapeutic agents for neurodegenerative diseases where protein aggregation and dysregulated RNA metabolism play central roles.
Relevance to metabolic disorders: Insights into glucose-driven O-GlcNAcylation provide a better understanding of how chronic hyperglycemia contributes to cellular dysfunction in diabetes and related complications.
Broader biomedical impact: By identifying candidate pathways and biomarkers sensitive to altered O-GlcNAcylation, this work offers new entry points for diagnostic and therapeutic development across multiple disease areas.
Overall, the study strengthens the link between metabolic state, protein modification, and disease mechanisms, paving the way toward more targeted and effective treatment strategies for conditions such as neurodegeneration and diabetes.
Potential impacts:
Advancement of therapeutic strategies: The results can inform the rational design and optimization of O-GlcNAcase inhibitors as potential therapeutic agents for neurodegenerative diseases where protein aggregation and dysregulated RNA metabolism play central roles.
Relevance to metabolic disorders: Insights into glucose-driven O-GlcNAcylation provide a better understanding of how chronic hyperglycemia contributes to cellular dysfunction in diabetes and related complications.
Broader biomedical impact: By identifying candidate pathways and biomarkers sensitive to altered O-GlcNAcylation, this work offers new entry points for diagnostic and therapeutic development across multiple disease areas.
Overall, the study strengthens the link between metabolic state, protein modification, and disease mechanisms, paving the way toward more targeted and effective treatment strategies for conditions such as neurodegeneration and diabetes.