PHASE I – WORK PLAN
Tooling for Measurement
Projects in Tooling for Measurement
Super-Resolution Ultra-Specific Imaging of Metabolites and Macromolecules by Second-Generation SRS Microscopy
This project aims to develop second-generation Stimulated Raman Scattering (SRS) microscopy for superresolution, ultra-specific imaging of metabolites and macromolecules in three-dimensional biological tissues. Current imaging techniques like fluorescence microscopy and mass spectrometry have limitations in resolution, specificity, and sample invasiveness. While SRS microscopy provides label-free chemical imaging with higher specificity, it lacks the resolution required for detailed analysis of nanoscopic metabolic activities in situ.
This project will combine advanced SRS nanoscopy techniques with new data analysis algorithms to create a platform capable of volumetric hyperspectral imaging at a resolution down to 14 nm. This will allow for unprecedented visualization of metabolic processes and molecular distributions in cells and tissues, providing deep insights into subcellular and tissue-level metabolic changes related to aging and diseases.
Transformative Mass Spectrometry Tools for Understanding Metabolic Changes in Aging
Aging is characterized by systemic metabolic changes that drive and reflect the progression of various age-related diseases, such as cancer, diabetes, and neurodegenerative disorders. Despite significant advancements in ‘omics technologies, a comprehensive understanding of the biochemical changes underlying aging remains incomplete. This project, led by experts in metabolism and mass spectrometry, aims to develop innovative approaches to analyze metabolic shifts associated with aging.
By integrating these approaches, the project aims to uncover the molecular underpinnings of metabolic dysregulation in aging, spatial disorganization of metabolism, and novel aging-associated biomolecules. The findings will provide actionable insights into the biochemistry of aging and offer potential pathways for therapeutic interventions targeting metabolic, inflammatory, and epigenetic processes.
Next Generation Mass Spectrometry-Based Proteomics for Characterization of Histone Post-Translational Modifications in Aging
The proposed research aims to develop next-generation mass spectrometry (MS) methodologies to characterize histone post-translational modifications (PTMs) in aging models, advancing our understanding of how chromatin modifications influence aging-related processes. Histone PTMs play a crucial role in regulating gene expression, chromatin structure, and cellular aging, but current techniques fall short in detecting low-abundance modifications. This project focuses on expanding the capabilities of both Bottom-Up and Top-Down MS approaches to enable in-depth analysis of histone PTMs, including combinatorial modifications in nucleosomes.
By optimizing liquid chromatography-mass spectrometry (LC-MS) workflows and incorporating advanced dissociation methods, such as electron-transfer dissociation (ETD) and ultraviolet photo dissociation (UVPD), we aim to accurately quantify over 500 histone PTMs in a single experiment. Furthermore, the Scout-MS approach will be developed for proteoform identification, targeting specific histone variants and modifications across the entire nucleosome. These innovative methods will be applied to aging related models to uncover chromatin alterations that drive age-associated changes, potentially revealing new therapeutic targets for age-related diseases. The project will generate an atlas of histone modifications and nucleosome conformational changes during aging, providing crucial insights into the epigenetic regulation of aging.
An Aging Biomarker for Living Cells: Development and Application to Real-Time Aging Analysis and Repair
Aging is a complex phenotype, resulting from the interaction of many genes, environmental factors, random events, and intricate cellular processes. Recent efforts have uncovered robust aging, longevity, and rejuvenation signatures by analyzing large-scale transcriptomic datasets across different tissues and organisms. However, omics technologies are destructive, and only offer us a snapshot into cellular age at a given moment in time.
Arguably, the most needed tool for aging biology is a biomarker that assesses aging in living cells. Such a biomarker could be used for real-time monitoring of aging under candidate interventions for promoting or counteracting aging, to assess which mechanisms of aging are affected by an intervention at specific times after the intervention is delivered, and as a measure to support closed-loop control of one or more aging-related processes within a system via real-time perturbation (perhaps empowered by machine learning). Indeed, a technology that captures the dynamic and highly interactive nature of the aging process at the single cell level could be the key to opening up the fundamental understanding and targeting of aging. Thus, it would enable a plethora of approaches, ranging from genetic and chemical screens, to longitudinal studies with live animals, to be coupled to real-time measurement of aging state.