Biology subjects

Masullo, L. A.

Publications and source records attributed to Masullo, L. A..

4 recordsLinked to original sources

bioRxiv2026

Distinct nanoscale membrane organizations of mucins and trans-sialidases in Trypanosoma cruzi

Trypanosoma cruzi, the causative agent of Chagas disease, relies on surface sialylation to evade host immunity and invade cells. This process is mediated by trans-sialidases (TS) and mucins, an enzyme-substrate pair anchored to distinct lipid environments. Yet, how these molecules are organized at the nanoscale in the parasites membrane remains unknown. Using dual-color super-resolution microscopy, we show that [~]60% of mucins and TS are segregated into nanoclusters ([~]100 nm) that rarely contact each other, distributed with a non-random separation distance, indicating that these abundant domains follow a specific order in the plasma membrane and are unlikely to serve as primary sites of sialylation. In contrast, the [~]40% fraction of non-clustered mucins and TS exhibits significantly shorter-than-random separation distances and appear ordered as in a shared fibrillar network. Additionally, we describe a distinct structural organization within these domains: mucins --residents of detergent-resistant domains (DRDs)--organize into high-molecular-weight complexes, whereas TS (which are excluded from DRDs) do not. This reveals an additional layer of membrane asymmetry and suggests a potential mechanism for domain-specific protein localization. Together, these findings uncover major principles of T. cruzi surface organization, with important implications for the regulation of host-parasite interactions.

microbiology↗

bioRxiv2026

Combinatorial DNA-PAINT

State-of-the-art super-resolution microscopy enables nanometer-resolution imaging of proteins, but its multiplexing capacity has been fundamentally constrained. Here, we present Combi-PAINT, a combinatorial DNA barcoding strategy based on DNA-PAINT, that facilitates superlinear scaling of multiplexing with the number of imaging rounds. Instead of assigning one unique docking strand per target, Combi-PAINT encodes targets as combinations of orthogonal sequences, allowing nonlinear scaling of target number with imaging rounds. Using just six imaging rounds, we resolve 41 targets in a field-of-view of 100 x 100 m2 with [~]2.5 nm localization precision and [~]90% decoding accuracy in under 30 minutes, representing the fastest sub-10 nm super-resolution microscopy acquisition to date. We benchmark decoding fidelity and demonstrate robust in situ performance in mammalian cells, achieving 97% decoding accuracy. Combi-PAINT is compatible with existing DNA-PAINT workflows and speed-enhancing techniques, offering a scalable, accessible platform for high-content, single-molecule spatial proteomics.

biophysics↗

bioRxiv2024

Structural diversity of Arc oligomers within the excitatory synapse

The Activity-Regulated Cytoskeleton-Associated protein (Arc), is pivotal to mediate plasticity responses in neuronal cells. In vitro studies suggest its ability to form high- and low-order oligomers, which are involved in neuronal trafficking. Despite its important functions, no direct observation of Arc oligomers in cells has been presented due to its highly regulated spatiotemporal expression, the small size of the structures, the lack of appropriate labelling strategies and the background associated to free diffusing cytosolic proteins. Here, we apply super resolution microscopy to observe Arc oligomeric states in cellular environment with focus on the excitatory synapse. In cells, we provide the first evidence of Arc high-order oligomers; we uncovered intermolecular interactions of Arc, its tendency to form liquid condensates and interaction with lipid bilayers. Arc high-order oligomers affect AMPA receptor surface levels. Together, our observations suggest a model by which Arc oligomerization mediates plasma membrane negative curvature favoring AMPA receptors endocytosis.

neuroscience↗

bioRxiv2023

Spatial proteomics in neurons at single-protein resolution

To fully understand biological processes and functions, it is necessary to reveal the molecular heterogeneity of cells and even subcellular assemblies by gaining access to the location and interaction of all biomolecules. The study of protein arrangements has seen significant advancements through super-resolution microscopy, but such methods are still far from reaching the multiplexing capacity of spatial proteomics. Here, we introduce Secondary label-based Unlimited Multiplexed DNA-PAINT (SUM-PAINT), a high-throughput imaging method capable of achieving virtually unlimited multiplexing at better than 15 nm spatial resolution. Using SUM-PAINT, we generated the most extensive multiprotein dataset to date at single-protein spatial resolution, comprising up to 30 distinct protein targets in parallel and adapted omics-inspired analysis workflows to explore these feature-rich datasets. Remarkably, by examining the multiplexed protein content of almost 900 individual synapses at single-protein resolution, we revealed the complexity of synaptic heterogeneity, ultimately leading to the discovery of a new synapse type. This work provides not only a feature-rich resource for researchers, but also an integrated data acquisition and analysis workflow for comprehensive spatial proteomics at single-protein resolution, paving the way for Localizomics.

neuroscience↗