Dennd3 functions as a guanine nucleotide exchange factor for small GTPase Rab12 in mouse embryonic fibroblasts

J Biol Chem. 2014 May 16;289(20):13986-95. doi: 10.1074/jbc.M113.546689. Epub 2014 Apr 9.


Small GTPase Rab12 regulates mTORC1 (mammalian target of rapamycin complex 1) activity and autophagy through controlling PAT4 (proton/amino acid transporter 4) trafficking from recycling endosomes to lysosomes, where PAT4 is degraded. However, the precise regulatory mechanism of the Rab12-mediated membrane trafficking pathway remained to be determined because a physiological Rab12-GEF (guanine nucleotide exchange factor) had yet to be identified. In this study we performed functional analyses of Dennd3, which has recently been shown to possess a GEF activity toward Rab12 in vitro. The results showed that knockdown of Dennd3 in mouse embryonic fibroblast cells caused an increase in the amount of PAT4 protein, the same as Rab12 knockdown did, and knockdown of Dennd3 and overexpression of Dennd3 were found to result in an increase and a decrease, respectively, in the intracellular amino acid concentration. Dennd3 overexpression was also found to reduce mTORC1 activity and promoted autophagy in a Rab12-dependent manner. Unexpectedly, however, Dennd3 knockdown had no effect on mTORC1 activity or autophagy despite increasing the intracellular amino acid concentration. Further study showed that Dennd3 knockdown reduced Akt activity, and the reduction in Akt activity is likely to have canceled out amino acid-induced mTORC1 activation through PAT4. These findings indicated that Dennd3 not only functions as a Rab12-GEF but also modulates Akt signaling in mouse embryonic fibroblast cells.

Keywords: Amino Acids Transport; Autophagy; Endosomes; G Proteins; Guanine Nucleotide Exchange Factor (GEF); Low Molecular Weight G Proteins; Membrane Trafficking; Rab Proteins; mTOR Complex (mTORC).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Transport Systems / metabolism
  • Amino Acids / metabolism
  • Animals
  • Autophagy
  • Cell Line
  • Fibroblasts / cytology
  • Fibroblasts / metabolism*
  • Gene Knockdown Techniques
  • Guanine Nucleotide Exchange Factors / deficiency
  • Guanine Nucleotide Exchange Factors / genetics
  • Guanine Nucleotide Exchange Factors / metabolism*
  • Intracellular Space / metabolism
  • Mechanistic Target of Rapamycin Complex 1
  • Mice
  • Multiprotein Complexes / metabolism
  • Proteolysis
  • Proto-Oncogene Proteins c-akt / metabolism
  • TOR Serine-Threonine Kinases / metabolism
  • rab GTP-Binding Proteins / metabolism*


  • Amino Acid Transport Systems
  • Amino Acids
  • Dennd3 protein, mouse
  • Guanine Nucleotide Exchange Factors
  • Multiprotein Complexes
  • PAT4 protein, mouse
  • Mechanistic Target of Rapamycin Complex 1
  • Proto-Oncogene Proteins c-akt
  • TOR Serine-Threonine Kinases
  • rab12 protein, mouse
  • rab GTP-Binding Proteins