The role of the tethering proteins p115 and GM130 in transport through the Golgi apparatus in vivo.
Biochemical data have shown that COPI-coated vesicles are tethered to Golgi membranes by a complex of at least three proteins: p115, giantin, and GM130. p115 binds to giantin on the vesicles and to GM130 on the membrane. We now examine the function of this tethering complex in vivo. Microinjection of an N-terminal peptide of GM130 or overexpression of GM130 lacking this N-terminal peptide inhibits the binding of p115 to Golgi membranes. Electron microscopic analysis of single microinjected cells shows that the number of COP-sized transport vesicles in the Golgi region increases substantially, suggesting that transport vesicles continue to bud but are less able to fuse. This was corroborated by quantitative immunofluorescence analysis, which showed that the intracellular transport of the VSV-G protein was significantly inhibited. Together, these data suggest that this tethering complex increases the efficiency with which transport vesicles fuse with their target membrane. They also provide support for a model of mitotic Golgi fragmentation in which the tethering complex is disrupted by mitotic phosphorylation of GM130.
Seemann
J
J
Cell Biology Laboratory, Imperial Cancer Research Fund, London WC2A 3PX, United Kingdom. joachim.seemann@yale.edu
Jokitalo
E J
EJ
Warren
G
G
D000818:Animals; D001324:Autoantigens; D001692:Biological Transport; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D006056:Golgi Apparatus / Q000378:metabolism* / Q000648:ultrastructure; D000077614:Golgi Matrix Proteins; D007399:Interphase; D007425:Intracellular Membranes / Q000378:metabolism / Q000648:ultrastructure; D008561:Membrane Fusion; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000008:administration & dosage / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008845:Microinjections; D008854:Microscopy, Electron; D010446:Peptide Fragments / Q000008:administration & dosage / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D017384:Sequence Deletion / Q000235:genetics; D014617:Vacuoles / Q000378:metabolism / Q000648:ultrastructure; D033921:Vesicular Transport Proteins; D014759:Viral Envelope Proteins / Q000378:metabolism
D001324:Autoantigens; D002352:Carrier Proteins; C028391:G protein, vesicular stomatitis virus; D000077614:Golgi Matrix Proteins; C097741:Golgin subfamily A member 2; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D010446:Peptide Fragments; D033921:Vesicular Transport Proteins; D014759:Viral Envelope Proteins; C091580:vesicular transport factor p115