Divergent functions of neuronal Rab11b in Ca2+-regulated versus constitutive exocytosis.
Using PC12 cells that express transfected human growth hormone (hGH) as a secreted reporter protein, we have searched for Rab proteins that function in exocytosis. Among the Rab proteins tested, we found that besides the previously described Rab3 proteins, only members of the Rab11 family (Rab11a, 11b, and 25) impaired Ca2+-induced exocytosis. Rab11b, which is enriched in brain, had the strongest effect. Consistent with a role in exocytosis, Rab11 and Rab3 proteins were colocalized with other vesicle proteins on secretory vesicles in PC12 cells and on mature synaptic vesicles in brain. Rab11b mutants that fix Rab11b in the GTP- or GDP-bound state both effectively inhibited Ca2+-induced exocytosis but seemed to act by distinct mechanisms: whereas GDP-bound Rab11b greatly stimulated constitutive secretion of hGH and depleted hGH stores in secretory vesicles, GTP-bound Rab11b only had a moderate effect on constitutive secretion and no effect on vesicular hGH stores. These results suggest that, consistent with a GTP-dependent regulation of Rab function, GDP-bound Rab11b indirectly inhibits Ca2+-triggered exocytosis by causing the loss of hGH from the PC12 cells, whereas GTP-bound Rab11b directly impairs Ca2+-triggered exocytosis. In contrast to neuroendocrine PC12 cells in which GTP- and GDP-bound Rab11b inhibited Ca2+-induced, but not constitutive, exocytosis, in non-neuronal cells GTP- and GDP-bound Rab11b inhibited constitutive exocytosis and caused an accumulation of cellular hGH. Viewed together, our data suggest that, in addition to other functions, Rab11 has a specific role in neuronal and neuroendocrine but not in non-neuronal cells as a GTP-dependent switch between regulated and constitutive secretory pathways.
Khvotchev
Mikhail V
MV
Center for Basic Neuroscience, Department of Molecular Genetics, and Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9111, USA.
Ren
Mindong
M
Takamori
Shigeo
S
Jahn
Reinhard
R
Südhof
Thomas C
TC
D000818:Animals; D002118:Calcium / Q000494:pharmacology / Q000502:physiology*; D002458:Cell Fractionation; D002460:Cell Line; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology*; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D019382:Human Growth Hormone / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D009154:Mutation; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D016716:PC12 Cells; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D022142:Secretory Vesicles / Q000378:metabolism; D013572:Synaptic Vesicles / Q000378:metabolism; D014162:Transfection; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D000099193:rab11 GTP-Binding Proteins
D002118:Calcium; D049452:Green Fluorescent Proteins; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D019382:Human Growth Hormone; D008164:Luminescent Proteins; D011188:Potassium; D011993:Recombinant Fusion Proteins; D020691:rab GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins