Stress-Induced Switch in Small Extracellular Vesicle Secretion: From Constitutive 'Torn Bag Mechanism' to Exocytosis

The biogenesis of small extracellular vesicles (sEVs) is only partially understood. Our recent findings provide evidence that a newly described sEV secretion pathway, the amphiectosome release followed by the sEV discharge by the 'torn bag mechanism' are present in all tested cell lines and mouse organs. Surprisingly, in in situ fixed steady-state cells, transmission electron microscopy did not reveal sEV release via exocytosis of multivesicular endosomes (MVEs). In the current study, we extended our previous analysis to additional mouse organs and confirmed the presence of secreted amphiectosomes in all of them. Furthermore, we investigated which parameters influence the activation of the distinct sEV release mechanisms in HEK cells. Our results show that under stress conditions (such as Ca²⁺ ionophore-induced membrane stress or metabolic stress, induced by serum starvation), exocytosis of MVEs is activated, while this process is absent in steady-state conditions. By silencing ATG5 (a key regulator of autophagy) and RAB27a (an essential small GTPase for MVE exocytosis), we selectively modulated these two mechanisms, respectively. Amphiectosome release depended on both autophagy and ATG5, while exocytosis of MVE was autophagy-independent but RAB27a-dependent. Our findings suggest that sEV release via the 'torn bag mechanism' is a general and essential secretion pathway in non-stressed, steady-state mammalian cells, while stress conditions induce the sEV release via MVE exocytosis.