SCAMP3 is essential for proper formation and function of neutrophil granules

Abstract

Host defense functions of neutrophils during infection critically depend on microbicidal and proteolytic proteins stored in primary, secondary and tertiary granules that are released into the phagosome or into the extracellular space upon degranulation. Granules are generated during granulopoiesis and impaired granule production or granule protein sorting has been linked to inefficient pathogen clearance resulting in recurrent bacterial and fungal infections. Here, we studied the role of the membrane protein secretory carrier associated membrane protein 3 (SCAMP3) for neutrophil defense functions. We generated Scamp3 knockout (KO) Hoxb8 cells and found that killing of Escherichia coli by Scamp3 KO Hoxb8 cell-derived neutrophils (dHoxb8 cells) was compromised as compared to control dHoxb8 cells in vitro. Mass spectrometric and Western blot analyses revealed a significant reduction of primary, secondary, and tertiary granule proteins in the genetic absence of Scamp3, resulting in a reduced overall granularity of these cells. Accordingly, degranulation was reduced in Scamp3 KO dHoxb8 cells compared to control dHoxb8 cells. Similarly, SCAMP3 deficiency in zebrafish resulted in reduced neutrophil granularity in comparison to wild-type animals. However, neutrophil migration towards sites of E. coli infection was unaffected in scamp3 KO zebrafish larvae. In summary, SCAMP3 represents an important novel player in granule equipment and degranulation, with key functions in neutrophil defense mechanisms during host-pathogen interactions in vitro. Brief summary sentence: The membrane protein secretory carrier associated membrane protein 3 (SCAMP3) is essential for proper protein equipment of neutrophil granules and Scamp3-deficient neutrophils have an impaired bacterial killing capacity in vitro.