A heterotrimeric G protein, G alpha i-3, on Golgi membranes regulates the secretion of a heparan sulfate proteoglycan in LLC-PK1 epithelial cells.
A heterotrimeric G alpha i subunit, alpha i-3, is localized on Golgi membranes in LLC-PK1 and NRK epithelial cells where it colocalizes with mannosidase II by immunofluorescence. The alpha i-3 was found to be localized on the cytoplasmic face of Golgi cisternae and it was distributed across the whole Golgi stack. The alpha i-3 subunit is found on isolated rat liver Golgi membranes by Western blotting and G alpha i-3 on the Golgi apparatus is ADP ribosylated by pertussis toxin. LLC-PK1 cells were stably transfected with G alpha i-3 on an MT-1, inducible promoter in order to overexpress alpha i-3 on Golgi membranes. The intracellular processing and constitutive secretion of the basement membrane heparan sulfate proteoglycan (HSPG) was measured in LLC-PK1 cells. Overexpression of alpha i-3 on Golgi membranes in transfected cells retarded the secretion of HSPG and accumulated precursors in the medial-trans-Golgi. This effect was reversed by treatment of cells with pertussis toxin which results in ADP-ribosylation and functional uncoupling of G alpha i-3 on Golgi membranes. These results provide evidence for a novel role for the pertussis toxin sensitive G alpha i-3 protein in Golgi trafficking of a constitutively secreted protein in epithelial cells.
Stow
J L
JL
Renal Unit, Massachusetts General Hospital, Boston.
de Almeida
J B
JB
Narula
N
N
Holtzman
E J
EJ
Ercolani
L
L
Ausiello
D A
DA
D000818:Animals; D001485:Basement Membrane / Q000378:metabolism; D002460:Cell Line; D011508:Chondroitin Sulfate Proteoglycans / Q000096:biosynthesis*; D004848:Epithelium / Q000502:physiology; D019204:GTP-Binding Proteins / Q000235:genetics / Q000502:physiology*; D015870:Gene Expression; D006056:Golgi Apparatus / Q000502:physiology* / Q000648:ultrastructure; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate / Q000096:biosynthesis*; D007124:Immunoenzyme Techniques; D007425:Intracellular Membranes / Q000502:physiology / Q000648:ultrastructure; D008099:Liver / Q000502:physiology; D046911:Macromolecular Substances; D008361:Mannosidases / Q000032:analysis; D008862:Microsomes, Liver / Q000502:physiology; D037342:Pertussis Toxin; D051381:Rats; D014162:Transfection; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D011508:Chondroitin Sulfate Proteoglycans; D019204:GTP-Binding Proteins; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate; D046911:Macromolecular Substances; D008361:Mannosidases; D037342:Pertussis Toxin; D010566:Virulence Factors, Bordetella; C038829:mannosyl-oligosaccharide 1,3 - 1,6-alpha-mannosidase