The rapid resealing of plasma membrane wounds is critical for cellular survival. Work supported by previous cycles of this grant showed that plasma membrane repair involves Ca2+-triggered exocytosis of lysosomes followed by massive endocytosis. Injury-induced release of lysosomal acid sphingomyelinase triggers a cholesterol/sphingolipid-dependent, clathrin-independent form of endocytosis, which involves caveolae or tubular invaginations that pinch off from the plasma membrane and carry lesions into cells for degradation. Clathrin-independent endocytosis has been described in many cell types, but there is considerable debate about how many independent pathways exist, and how they are initiated. By demonstrating that plasma membrane injury, Ca2+ influx and secretion of lysosomal acid sphingomyelinase trigger clathrin-independent endocytosis, our results have introduced much needed clarity to this field. The studies we now propose provide a unique opportunity for understanding how clathrin-independent endocytosis is regulated by plasma membrane injury, and how it promotes wound removal. Strikingly, our recent studies in B lymphocytes suggest that endocytosis-dependent plasma membrane repair and BCR-mediated B cell activation interfere with each other because of competition for lipid rafts, the cholesterol/sphingolipid-enriched plasma membrane microdomains that play a central role in clathrin-independent endocytosis and in BCR signaling and internalization after antigen capture. To understand the physiological impact of these findings, we will pursue two specific aims: 1) Characterize the injury-induced clathrin-independent form of endocytosis that promotes plasma membrane repair; 2) Examine the impact of plasma membrane wounding and repair on the regulation of B cell activation. These studies will significantly advance our understanding of clathrin-independent endocytosis and its role in plasma membrane resealing, and demonstrate how plasma membrane injury and repair regulate the function of B lymphocytes, cells that play an essential role in immune protection against infectious agents.

Public Health Relevance

This work is focused on understanding how cells remove wounds from their membrane, so it is relevant to the development of treatments for diseases such as muscular dystrophy that are caused by frequent injury and defective membrane repair. This project will also investigate how internalization of plasma membrane wounds affects the function of B lymphocytes, cells that produce antibodies and thus play a critical role in immune responses. By determining how internalization of wounded membrane influences the ability of B cells to respond to pathogens, our project will create new opportunities for a better understanding and potential modulation of immune responses.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Membrane Biology and Protein Processing Study Section (MBPP)
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Flicker, Paula F
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University of Maryland College Park
Anatomy/Cell Biology
Earth Sciences/Resources
College Park
United States
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Castro-Gomes, Thiago; Corrotte, Matthias; Tam, Christina et al. (2016) Plasma Membrane Repair Is Regulated Extracellularly by Proteases Released from Lysosomes. PLoS One 11:e0152583
Corrotte, M; Castro-Gomes, T; Koushik, A B et al. (2015) Approaches for plasma membrane wounding and assessment of lysosome-mediated repair responses. Methods Cell Biol 126:139-58
Miller, Heather; Castro-Gomes, Thiago; Corrotte, Matthias et al. (2015) Lipid raft-dependent plasma membrane repair interferes with the activation of B lymphocytes. J Cell Biol 211:1193-205
Fernandes, Maria Cecilia; Corrotte, Matthias; Miguel, Danilo C et al. (2015) The exocyst is required for trypanosome invasion and the repair of mechanical plasma membrane wounds. J Cell Sci 128:27-32
Andrews, N W; Corrotte, M; Castro-Gomes, T (2015) Above the fray: Surface remodeling by secreted lysosomal enzymes leads to endocytosis-mediated plasma membrane repair. Semin Cell Dev Biol 45:10-7
Andrews, N W; Perez, F (2015) The plasma membrane repair shop: Fixing the damage. Semin Cell Dev Biol 45:1
Castro-Gomes, T; Koushik, A B; Andrews, N W (2014) ESCRT: nipping the wound in the bud? Trends Biochem Sci 39:307-9
Andrews, Norma W; Almeida, Patricia E; Corrotte, Matthias (2014) Damage control: cellular mechanisms of plasma membrane repair. Trends Cell Biol 24:734-42
Hissa, Barbara; Pontes, Bruno; Roma, Paula Magda S et al. (2013) Membrane cholesterol removal changes mechanical properties of cells and induces secretion of a specific pool of lysosomes. PLoS One 8:e82988
Flannery, Andrew R; Renberg, Rebecca L; Andrews, Norma W (2013) Pathways of iron acquisition and utilization in Leishmania. Curr Opin Microbiol 16:716-21

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