PROJECT SUMMARY Bacterial coinfections have been the leading cause of severe illness and mortality during past Influenza pandemics and current seasonal endemics. Streptococcus pneumoniae is the most common etiological agent reported during Influenza-associated secondary infections. Unfortunately, the epithelial and immunological mechanisms resulting in increased pneumococcal invasion, persistence, and dissemination in the Influenza aftermath remain poorly understood. Preliminary data suggest that Microfold (M) cells arise in the lung epithelium during the resolution phase of Influenza-associated inflammation when susceptibility to pneumococcal infections is maximum. Highly endocytic intestinal M cells play a crucial role in gut immune surveillance by directly delivering luminally captured antigens to underlying dendritic cells, eliciting antigen- specific mucosal immunity. However, M cells in the gut are occasionally hijacked and serve as entry portals for pathogenic dissemination into the host. Having been only recently identified, our knowledge of lung M cell functions in pulmonary infection and immunity is scant. Our single-cell transcriptomic analysis of the murine airway epithelium reveals M cell-specific expression of novel transcytosis receptors, chemokines, and regulators of endocytic trafficking with immunomodulatory properties. This proposal aims to define the functional role of lung M cells in predisposition to influenza-associated pneumococcal invasion and ensuing pneumonia and sepsis. We further aim to identify the intracellular trafficking mechanisms during pneumococcal transcytosis across airway M cells. The proposed research goals will be achieved by employing in vitro and in vivo models of M cell induction in the lower airways coupled with genetically engineered mouse models for M cell ablation and innovative live imaging platforms. This study will shed light on the M cell-driven mechanisms promoting invasive pneumococcal disease and will inform the development of ingenious therapeutic strategies to target epithelial entry portals, thus preventing bacterial superinfections in the background of viral infections. The proposed research plan will be executed at the MGH Center for Regenerative Medicine (CRM), Boston, under the mentorship of Dr. Jayaraj Rajagopal, an expert in the field of airway epithelial biology and lung regeneration. My career objective is to become a tenure-track faculty pioneering and simultaneously training next-generation scientists in the field of lung infections and immunity. To accomplish my career goals, I have put together a comprehensive training plan to enhance the overall skill sets required to establish myself as a successful independent investigator. To ensure timely progress toward fulfilling my rigorous research plan and career goals, I have gathered an expert advisory committee comprising Dr. Dennis Brown, Dr. Jatin Vyas, and Dr. David Scadden, with whom I will regularly discuss my research progre...