398.4 - Exosomes as Key Regulators of Neutrophil Chemotaxis

Carole Parent (University of Michigan, University of Michigan, University of Michigan, University of Michigan), Subhash Arya (University of Michigan, University of Michigan), Song Chen (University of Michigan), Fatima Javed (University of Michigan)

Neutrophils, the most abundant leukocytes in human blood, are the first line of defense at sites of injury or infection. Upon activation by primary chemoattractants such as the formyl peptide N-Formyl-methionine-leucyl-phenylalanine (fMLF), neutrophils secrete the secondary chemoattractant leukotriene B4 (LTB4) which serves to relay primary chemotactic signals and increase the range and persistence of migration. LTB4 is synthesized from arachidonic acid (AA) through the sequential action of the 5-lipoxygenase (5-LO), its associated activating protein (FLAP) and leukotriene A4 hydrolase (LTA4H). We previously demonstrated that LTB4 and its synthesizing enzymes are packaged and released in extracellular vesicles called exosomes. Conventional exosome biogenesis originates from the plasma membrane. However, the LTB4 synthesizing machinery resides on the nuclear envelope (NE). We found that the localized activation of membrane curvature inducing proteins facilitates the selective sorting of the LTB4-synthesis machinery on the NE and its packaging within NE-derived exosomes. We show that chemoattractant stimulation leads to the generation of 5-LO- and FLAP-positive NE buds and cytosolic vesicles, which are also positive for the inner nuclear membrane protein lamin B receptor (LBR). Furthermore, we found that the generation of ceramide from sphingomyelin through the action of the neutral sphingomyelinase 1 (nSMase1) is essential for the generation of these NE buds and cytosolic vesicles by generating ordered lipid microdomains on the NE. Together, our findings show that the spatiotemporal orchestration of chemoattractant-initiated events leads to the recruitment of the LTB4 biogenesis machinery on the NE followed by NE-budding and subsequent release of cytosolic vesicles containing exosomes. We envision that this novel mechanism will represent an important means of exosome biogenesis for the release of NE-associated components in other cells, including immune cells and invasive cancer cells.

Research supported by funds from the University of Michigan School of Medicine and NIH R01AI152517 to CAP.