Distribution of Signal Peptides in Microvesicles from Activated Macrophage Cells

Extracellular vesicles, such as microvesicles (LEV) and exosomes (SEV), play an important role in intercellular signaling by encapsulating functional molecules and delivering them to specific cells. Recent studies showed that signal peptides (SPs), which are derived from sequences at the N-terminal of newly synthesized proteins, exhibited biological activity in the extracellular fluid. We previously reported that SPs were secreted into the extracellular fluid via SEV; however, it remains unclear whether the release of SPs occurs via LEV. In the present study, we demonstrated that SP fragments from human placental secreted alkaline phosphatase (SEAP) were present in LEV as well as SEV released from RAW-Blue cells, which stably express an NF-& kappa;B-inducible SEAP reporter. When RAW-Blue cells were treated with LPS at 0-10,000 ng/mL, SEAP SP fragments per particle were more abundant in LEV than in SEV, with fragments in LEV and SEV reaching a maximum at 1000 and 100 ng/mL, respectively. The content of SEAP SP fragments in LEV from IFN & gamma;-stimulated RAW-Blue cells was higher than those from TNF & alpha;-stimulated cells, whereas that in SEV from TNF & alpha;-stimulated RAW-Blue cells was higher than those from IFN & gamma;-stimulated cells. Moreover, the content of SEAP SP fragments in LEV and SEV decreased in the presence of W13, a calmodulin inhibitor. Collectively, these results indicate that the transportation of SP fragments to extracellular vesicles was changed by cellular activation, and calmodulin was involved in their transportation to LEV and SEV.

Automated summary

Extracellular vesicles, including microvesicles and exosomes, are involved in intercellular signaling by delivering functional molecules. This study demonstrated the presence of signal peptide (SP) fragments from human placental secreted alkaline phosphatase (SEAP) in both microvesicles (LEV) and exosomes (SEV). The content of SEAP SP fragments in LEV and SEV was influenced by cellular activation, and calmodulin was found to be involved in their transportation.