Spatial partitioning of secretory cargo from Golgi resident proteins in live cells
1 Light Microscopy Group,European Molecular Biology Laboratory(EMBL),Meyerhofstrabe Heidelberg, Germany
2 Cell Biology and Biophysics Programme European Molecular Biology Laboratory (EMBL) Meyerhofstraβe, Heidelberg, Germany
3 Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraβe 108, Dresden, Germany
4 Massachusetts General Hospital Cancer Research Center, Charlestown, Massachusetts, USA
BMC Cell Biology 2001, 2:19 doi:10.1186/1471-2121-2-19Published: 10 October 2001
To maintain organelle integrity, resident proteins must segregate from itinerant cargo during secretory transport. However, Golgi resident enzymes must have intimate access to secretory cargo in order to carry out glycosylation reactions. The amount of cargo and associated membrane may be significant compared to the amount of Golgi membrane and resident protein, but upon Golgi exit, cargo and resident are efficiently sorted. How this occurs in live cells is not known.
We observed partitioning of the fluorescent Golgi resident T2-CFP and fluorescent cargo proteins VSVG3-YFP or VSVG3-SP-YFP upon Golgi exit after a synchronous pulse of cargo was released from the ER. Golgi elements remained stable in overall size, shape and relative position as cargo emptied. Cargo segregated from resident rapidly by blebbing into micron-sized domains that contained little or no detectable resident protein and that appeared to be continuous with the parent Golgi element. Post-Golgi transport carriers (TCs) exited repeatedly from these domains. Alternatively, entire cargo domains exited Golgi elements, forming large TCs that fused directly with the plasma membrane. However, domain formation did not appear to be an absolute prerequisite for TC exit, since TCs also exited directly from Golgi elements in the absence of large domains. Quantitative cargo-specific photobleaching experiments revealed transfer of cargo between Golgi regions, but no discrete intra-Golgi TCs were observed.
Our results establish domain formation via rapid lateral partitioning as a general cellular strategy for segregating different transmembrane proteins along the secretory pathway and provide a framework for consideration of molecular mechanisms of secretory transport.