Localization and targeting of SCG10 to the trans-Golgi apparatus and growth cone vesicles.
SCG10 is a membrane-associated, microtubule-destabilizing protein of neuronal growth cones. Using immunoelectron microscopy, we show that in the developing cortex of mice, SCG10 is specifically localized to the trans face Golgi complex and apparently associated with vesicular structures in putative growth cones. Consistent with this, subcellular fractionation of rat forebrain extracts demonstrates that the protein is enriched in the fractions containing the Golgi apparatus and growth cone particles. In isolated growth cone particles, SCG10 was found to be particularly concentrated in the growth cone vesicle fraction. To evaluate the molecular determinants of the specific targeting of SCG10 to growth cones, we have transfected PC12 cells and primary neurons in culture with mutant and fusion cDNA constructs. Deletion of the amino-terminal domain or mutations within this domain that prevented palmitoylation at cysteines 22 and 24 abolished Golgi localization as well as growth cone targeting, suggesting that palmitoylation of the amino-terminal domain is a necessary signal for Golgi sorting and possibly transport of SCG10 to growth cones. Fusion proteins consisting of the amino-terminal domain of SCG10 and the cytosolic proteins stathmin or glutathione-S-transferase colocalized with a Golgi marker, alpha-mannosidase II, and accumulated in growth cones of both axons and dendrites. These results reveal a novel axonal/dendritic growth cone targeting sequence that involves palmitoylation.
Lutjens
R
R
Institut de Biologie Cellulaire et de Morphologie, University of Lausanne, Switzerland.
Igarashi
M
M
Pellier
V
V
Blasey
H
H
Di Paolo
G
G
Ruchti
E
E
Pfulg
C
C
Staple
J K
JK
Catsicas
S
S
Grenningloh
G
G
D000818:Animals; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D003545:Cysteine / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D017353:Gene Deletion; D006056:Golgi Apparatus / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D020439:Growth Cones / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D007425:Intracellular Membranes / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D051379:Mice; D008815:Mice, Inbred Strains; D016253:Microscopy, Immunoelectron; D008868:Microtubule Proteins; D016296:Mutagenesis / Q000502:physiology; D009414:Nerve Growth Factors / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*; D009419:Nerve Tissue Proteins / Q000032:analysis; D016716:PC12 Cells; D019308:Palmitic Acid / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics; D017434:Protein Structure, Tertiary; D051381:Rats; D050777:Stathmin; D013347:Subcellular Fractions / Q000737:chemistry; D016708:Synaptophysin / Q000032:analysis; D050825:Synaptosomal-Associated Protein 25; D014162:Transfection
D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D008868:Microtubule Proteins; D009414:Nerve Growth Factors; D009419:Nerve Tissue Proteins; D021382:Protein Sorting Signals; C494209:Snap25 protein, mouse; C494210:Snap25 protein, rat; D050777:Stathmin; C512849:Stmn2 protein, mouse; C512848:Stmn2 protein, rat; D016708:Synaptophysin; D050825:Synaptosomal-Associated Protein 25; D019308:Palmitic Acid; D003545:Cysteine