Distribution of G-proteins in rat liver plasma-membrane domains and endocytic pathways.
1. The distribution of the alpha- and beta-subunits of nucleotide-binding G-proteins among rat liver sinusoidal, lateral and canalicular plasma membranes, endosomes, Golgi membranes and lysosomes was investigated. 2. Pertussis-toxin-catalysed ADP-ribosylation identified a 41 kDa inhibitory alpha-subunit in all liver plasma-membrane functional domains as well as in endosomes. An antibody to a synthetic peptide corresponding to a C-terminal sequence of the inhibitory alpha-subunit also identified the 41 kDa polypeptide in all plasma-membrane domains, in 'early' and 'late' endosomes and in Golgi membranes; this polypeptide was not detected in lysosomes. The antibody-binding studies showed that bile-canalicular plasma membranes had the highest content of the inhibitory alpha-subunit. 3. Immunofluorescent microscopy confirmed the presence of the inhibitory alpha-subunit in all regions of the hepatocyte's cell surface. 4. An antibody recognizing the beta-subunit showed that a 36 kDa polypeptide was present in all plasma membranes and in 'early' and 'late' endosomes; it was not detected in lysosomes. The relative distribution among the fractions of this polypeptide was similar to the distribution of the inhibitory alpha-subunit. 5. The presence of high levels of the G-protein inhibitory alpha-subunit in bile-canalicular plasma membranes was confirmed by demonstration of its co-fractionation with marker enzymes in Nycodenz gradients and by free-flow electrophoresis. The significance of this location is discussed.
Ali
N
N
National Institute for Medical Research, Mill Hill, London, U.K.
Milligan
G
G
Evans
W H
WH
D000818:Animals; D001648:Bile Canaliculi / Q000032:analysis; D015153:Blotting, Western; D002462:Cell Membrane / Q000032:analysis; D004705:Endocytosis; D019204:GTP-Binding Proteins / Q000032:analysis*; D008099:Liver / Q000032:analysis* / Q000648:ultrastructure; D008247:Lysosomes / Q000032:analysis; D008297:Male; D015388:Organelles / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains
D019204:GTP-Binding Proteins