TBC1D12 is a novel Rab11-binding protein that modulates neurite outgrowth of PC12 cells

PLoS One. 2017 Apr 6;12(4):e0174883. doi: 10.1371/journal.pone.0174883. eCollection 2017.

Abstract

Recycling endosomes are generally thought to play a central role in endocytic recycling, but recent evidence has indicated that they also participate in other cellular events, including cytokinesis, autophagy, and neurite outgrowth. Rab small GTPases are key regulators in membrane trafficking, and although several Rab isoforms, e.g., Rab11, have been shown to regulate recycling endosomal trafficking, the precise mechanism by which these Rabs regulate recycling endosomes is not fully understood. In this study, we focused on a Rab-GTPase-activating protein (Rab-GAP), one of the key regulators of Rabs, and comprehensively screened 43 mammalian Tre-2/Bub2/Cdc16 (TBC)/Rab-GAP-domain-containing proteins (TBC proteins) for proteins that specifically localize on recycling endosomes in mouse embryonic fibroblasts (MEFs). Four of the 43 mammalian TBC proteins screened, i.e., TBC1D11, TBC1D12, TBC1D14, and EVI5, were found to colocalize well with transferrin receptor, a well-known recycling endosome marker. We further investigated the biochemical properties of TBC1D12, a previously uncharacterized TBC protein. The results showed that TBC1D12 interacted with active Rab11 through its middle region and that it did not display Rab11-GAP activity in vitro. The recycling endosomal localization of TBC1D12 was found to depend on the expression of Rab11. We also found that TBC1D12 expression had no effect on common Rab11-dependent cellular events, e.g., transferrin recycling, in MEFs and that it promoted neurite outgrowth, a specialized Rab11-dependent cellular event, of PC12 cells independently of its GAP activity. These findings indicated that TBC1D12 is a novel Rab11-binding protein that modulates neurite outgrowth of PC12 cells.

MeSH terms

  • Animals
  • Cells, Cultured
  • GTP-Binding Proteins / metabolism*
  • GTPase-Activating Proteins / genetics
  • GTPase-Activating Proteins / metabolism
  • GTPase-Activating Proteins / physiology*
  • Gene Knockdown Techniques
  • Mice
  • Neuronal Outgrowth / physiology*
  • PC12 Cells
  • Rats

Substances

  • GTPase-Activating Proteins
  • TBC1D12 protein, rat
  • Wdr44 protein, rat
  • GTP-Binding Proteins

Grants and funding

This work was supported by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, and Technology (MEXT) of Japan (grant number 15H04367 to M.F.). There was no additional external funding received for this study.