Abstract
Rab39a has pleiotropic functions in phagosome maturation, inflammatory activation and neuritogenesis. Here, we characterized Rab39a function in membrane trafficking of phagocytosis and autophagy induction in macrophages. Rab39a localized to the periphery of LAMP2-positive vesicles and showed the similar kinetics on the phagosome to that of LAMP1. The depletion of Rab39a did not influence the localization of LAMP2 to the phagosome, but it augments the autophagosome formation and LC3 processing by lipopolysaccharide (LPS) stimulation. The augmentation of autophagosome formation in Rab39a-knockdown macrophages was suppressed by Atg5 depletion or an inhibitor for phosphatidylinostol 3-kinase (PI3K). Immunoprecipitation analysis revealed that Rab39a interacts with PI3K and that the amino acid residues from 34th to 41st in Rab39a were indispensable for this interaction. These results suggest that Rab39a negatively regulates the LPS-induced autophagy in macrophages.
Highlights
Rab GTPases localize to specific subcellular organelles and regulate various membrane trafficking [1,2]
Rab39a localizes to lysosomes To characterize the localization of Rab39a in macrophages, Raw264.7 macrophages were transfected with EGFP-Rab39a and immunostained with anti-LAMP2 antibody
By Fluorescence recovery after photobleaching (FRAP) analysis, we can assess the state of activation of Rab39a on the membrane, because the active GTP-bound forms of Rab GTPases are stable on the membrane while the inactive GDPbound forms release from the membrane to the cytosol
Summary
Rab GTPases localize to specific subcellular organelles and regulate various membrane trafficking [1,2]. We screened and identified Rab GTPases that regulate phagosome maturation in macrophages [7]. One of these Rab GTPases, Rab39a was involved in phagosomal acidification [7]. Rab39a has been demonstrated to be involved in the regulation of Caspase-1 activity [8] and the differentiation of neuron cells [9]. These results together indicate that Rab39a has pleiotropic functions in phagosome maturation, inflammatory activation and neuritogenesis
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