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Golvesin‐GFP fusions as distinct markers for Golgi and post‐Golgi vesicles in Dictyostelium cells
Author(s) -
Schneider Natalie,
Schwartz JeanMarc,
Köhler Jana,
Becker Michael,
Schwarz Heinz,
Gerisch Günther
Publication year - 2000
Publication title -
biology of the cell
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.543
H-Index - 85
eISSN - 1768-322X
pISSN - 0248-4900
DOI - 10.1016/s0248-4900(00)01102-3
Subject(s) - golgi apparatus , microbiology and biotechnology , endosome , biology , dictyostelium , vesicle , organelle , green fluorescent protein , vacuole , brefeldin a , transport protein , cell plate , cytokinesis , cytoplasm , biochemistry , cell division , endoplasmic reticulum , membrane , cell , gene , intracellular
Golvesin is a new protein associated with membranes of the Golgi apparatus and post‐Golgi vesicles in Dictyostelium cells. An internal hydrophobic sequence of 24 amino‐acid residues is responsible for anchoring golvesin to the membranes of these organelles. In an attempt to visualize organelle dynamics in vivo, we have used specific antibody and other labels to localize golvesin—green fluorescent protein (GFP) constructs to different cellular compartments. With a GFP tag at its N‐terminus, golvesin shows the same localization as the untagged protein. It is transferred to two post‐Golgi compartments, the endosomal and contractile vacuole systems. Endosomes are decorated with GFP—golvesin within less than 10 min of their internalisation, and keep the label during the acidic phase of the pathway. Blockage of the C‐terminus with GFP causes entrapment of the protein in the Golgi apparatus, indicating that a free C‐terminus is required for transfer of golvesin to any of the post‐Golgi compartments. The C‐terminally tagged golvesin proved to be a reliable Golgi marker in Dictyostelium cells revealing protrusion of Golgi tubules at peak velocities of 3 to 4 μm·s –1 . The fusion protein is retained in Golgi vesicles during mitosis, visualizing Golgi disassembly and reorganization in line with cytokinesis.