Our research focus

We aim to unravel the intricate intracellular processes governing plasma membrane rearrangements and actomyosin cytoskeleton remodeling, which are pivotal events for membrane resealing and thus constitute critical components of cell survival and defense against infections. By elucidating these mechanisms, we seek to enrich our comprehension of how host cells endure infectious damage, thereby laying the groundwork for innovative therapeutic strategies to combat infections and promote tissue repair.

The loss of plasma membrane integrity triggers uncontrolled exchanges between the intracellular and extracellular environments, precipitating the disruption of cellular homeostasis. Notably, our research has revealed that the massive influx of calcium through the toxin-based pores induces endoplasmic reticulum (ER) disruption and prompts the recruitment of ER-derived vacuoles to the cell cortex. This process occurs concurrently with profound remodeling of the cortical actomyosin cytoskeleton and extensive plasma membrane blebbing. Importantly, we have established a correlation between plasma membrane blebbing, actomyosin cytoskeleton remodeling, and the resolution of damage, demonstrating their regulation by ER proteins.

Based on available data, we postulate that the recruitment of ER-derived vacuoles to the cell cortex serves a dual purpose: firstly, as a source of new membrane for damage repair, and secondly, to polarize essential intracellular machinery to the site of injury. To further investigate our hypothesis, we employ a multifaceted approach encompassing proximity-labeling proteomics, live-cell imaging, and advanced high-resolution and combinatory electron microscopy techniques.

To induce damage in host cell lines and deeply investigate common repair mechanisms, we employ a variety of damaging agents, including:

1. Purified recombinant pore-forming toxins, such as PLY (Pneumolysin), LLO (Listeriolysin O), and PFO (Perfringolysin O);
2. Bacterial pathogens known to produce pore-forming toxins, including Streptococcus pneumoniae, Listeria monocytogenes, and Clostridium perfringens;
3. Influenza A virus, which causes significant damage to epithelial lung cells.

In our laboratory, we employ a diverse range of cellular models for our research. These include commercially available epithelial cell lines, genetically modified cell lines, 3D cell models cultured at the air-liquid interface, and stem cell-derived 3D cell models. To evaluate infection outcomes, we use zebrafish or C. elegans infection models.

We are currently addressing several key issues, including:

1. Characterizing novel repair machineries, with a specific emphasis on calcium-responsive proteins;
2. Resolving the cellular ultrastructure in the vicinity of the pore and the plasma membrane blebbing, to uncover new cellular compartments involved in the repair process;
3. Elucidating the molecular intricacies of the crosstalk between the endoplasmic reticulum (ER) and cytoskeletal components;
4. Investigating the spatiotemporal regulation of repair events.

Ongoing projects

Photo Credits: Joana Pereira