Pancreatitis is characterized by exocrine pancreatic insufficiency due to malfunctioning of the ion secretion homeostasis within the pancreatic ductal epithelial cells. Proper functioning of the cAMP-mediated protein kinase A-regulated cystic fibrosis transmembrane conductance regulator (CFTR) HCO3-/Cl- channel is primordial to control the viscosity of the pancreatic juice responsible for adequate delivery of digestive enzymes into the duodenum. cAMP is produced from ATP through the activity of the adenylyl cyclases; some of which are regulated by Ca2+. Cytoplasmic Ca2+ fluxes are generated by Ca2+ release from the ER store into the cytoplasmic space and subsequent uptake –through store-operated Ca2+ entry (SOCE)– from the extracellular space into the cell.
Confining production of the ubiquitous second messenger cAMP to discrete subcellular locations allows for unique spatiotemporal functional responses. Yet, the factors that determine the compartmentation of this diffusible signaling molecule are not understood. Although the fluid mosaic model originally proposed that signal transduction occurs through random interactions between proteins diffusing freely throughout the plasma membrane (PM), it is now established that the PM is segregated into distinct microdomains (rafts) where signaling proteins are concentrated.
In the current proposal, we first want to analyze what role lipid raft and non-raft membrane domains play in compartmentation of Ca2+-induced cAMP signaling and subsequent CFTR activation in 2D cell culture. We will combine state-of-the-art molecular biological and imaging approaches such as FRET-based cAMP sensors and cAMP chelators to monitor and manipulate SOCE-induced spatiotemporal changes in cAMP required for CFTR activation. Through targeted enzyme-induced lipid raft modulations, we will analyze the involvement of these microdomains on SOCE-induced cAMP-mediated CFTR activation. Next, we aim to confirm and analyze the physiological relevance of the obtained findings in polarized epithelial cells and human 3-dimensional (3D) pancreatic ductal epithelial organoid cultures. The obtained insights will provide a better understanding of the functional role of lipid raft and non-raft membrane domains in compartmentation of Ca2+-induced cAMP signaling in polarized epithelial cells and might provide valuable clues for the development of efficient therapeutic strategies to alleviate pancreatitis.
előírt nyelvtudás: angol felvehető hallgatók száma: 1
Jelentkezési határidő: 2022-06-30
2024. IV. 17. ODT ülés Az ODT következő ülésére 2024. június 14-én, pénteken 10.00 órakor kerül sor a Semmelweis Egyetem Szenátusi termében (Bp. Üllői út 26. I. emelet).
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