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Visualization of the dynamics of synaptic vesicle and plasma membrane proteins in living axons.

Nakata T, Terada S, Hirokawa N - J. Cell Biol. (1998)

Bottom Line: We have successfully visualized the transporting vesicles of plasma membrane proteins, synaptic vesicle proteins, and the trans-Golgi network residual proteins in living axons at high resolution using laser scan microscopy of green fluorescent protein-tagged proteins after photobleaching.We found that all of these proteins are transported by tubulovesicular organelles of various sizes and shapes that circulate within axons from branch to branch and switch the direction of movement.These organelles are distinct from the endosomal compartments and constitute a new entity of membrane organelles that mediate the transport of newly synthesized proteins from the trans-Golgi network to the plasma membrane.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Anatomy, Graduate School of Medicine, University of Tokyo, Hongo, Tokyo, Japan, 113.

ABSTRACT
Newly synthesized membrane proteins are transported by fast axonal flow to their targets such as the plasma membrane and synaptic vesicles. However, their transporting vesicles have not yet been identified. We have successfully visualized the transporting vesicles of plasma membrane proteins, synaptic vesicle proteins, and the trans-Golgi network residual proteins in living axons at high resolution using laser scan microscopy of green fluorescent protein-tagged proteins after photobleaching. We found that all of these proteins are transported by tubulovesicular organelles of various sizes and shapes that circulate within axons from branch to branch and switch the direction of movement. These organelles are distinct from the endosomal compartments and constitute a new entity of membrane organelles that mediate the transport of newly synthesized proteins from the trans-Golgi network to the plasma membrane.

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Generation of a transporting vesicle in the cell body.  (a) The cell was transfected with adenovirus vector carrying GAP-43–GFP chimeric DNA. A bright label within the cell body was  observed. (b) Time series imaging of a part of the cell body in a.  A bright punctuate staining emerged from static intense GFP-staining in the cell body (2nd frame). From there, a tubular vesicle protruded at the speed of fast axonal transport (2nd–4th  frames), separated from the static intense GFP-staining (5th  frame), and proceeded into the axon (6th and 7th frames). Diamonds indicate the distal edge of the tubule. Intervals between  frames are 3.45 s. Bar, 5μm.
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Figure 6: Generation of a transporting vesicle in the cell body. (a) The cell was transfected with adenovirus vector carrying GAP-43–GFP chimeric DNA. A bright label within the cell body was observed. (b) Time series imaging of a part of the cell body in a. A bright punctuate staining emerged from static intense GFP-staining in the cell body (2nd frame). From there, a tubular vesicle protruded at the speed of fast axonal transport (2nd–4th frames), separated from the static intense GFP-staining (5th frame), and proceeded into the axon (6th and 7th frames). Diamonds indicate the distal edge of the tubule. Intervals between frames are 3.45 s. Bar, 5μm.

Mentions: Using the above approach, we investigated the generation of transporting vesicles in the cell body (Fig. 6). We found that tubules arose, protruded, and pinched off from the bright staining of GAP-43–GFP within the cell body, the distribution of which was similar to that of TGN-38 (Fig. 4). The vesicles moved at the speed of fast axonal transport as soon as they were generated in the cell body, and moved directly into the axon.


Visualization of the dynamics of synaptic vesicle and plasma membrane proteins in living axons.

Nakata T, Terada S, Hirokawa N - J. Cell Biol. (1998)

Generation of a transporting vesicle in the cell body.  (a) The cell was transfected with adenovirus vector carrying GAP-43–GFP chimeric DNA. A bright label within the cell body was  observed. (b) Time series imaging of a part of the cell body in a.  A bright punctuate staining emerged from static intense GFP-staining in the cell body (2nd frame). From there, a tubular vesicle protruded at the speed of fast axonal transport (2nd–4th  frames), separated from the static intense GFP-staining (5th  frame), and proceeded into the axon (6th and 7th frames). Diamonds indicate the distal edge of the tubule. Intervals between  frames are 3.45 s. Bar, 5μm.
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Related In: Results  -  Collection

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Figure 6: Generation of a transporting vesicle in the cell body. (a) The cell was transfected with adenovirus vector carrying GAP-43–GFP chimeric DNA. A bright label within the cell body was observed. (b) Time series imaging of a part of the cell body in a. A bright punctuate staining emerged from static intense GFP-staining in the cell body (2nd frame). From there, a tubular vesicle protruded at the speed of fast axonal transport (2nd–4th frames), separated from the static intense GFP-staining (5th frame), and proceeded into the axon (6th and 7th frames). Diamonds indicate the distal edge of the tubule. Intervals between frames are 3.45 s. Bar, 5μm.
Mentions: Using the above approach, we investigated the generation of transporting vesicles in the cell body (Fig. 6). We found that tubules arose, protruded, and pinched off from the bright staining of GAP-43–GFP within the cell body, the distribution of which was similar to that of TGN-38 (Fig. 4). The vesicles moved at the speed of fast axonal transport as soon as they were generated in the cell body, and moved directly into the axon.

Bottom Line: We have successfully visualized the transporting vesicles of plasma membrane proteins, synaptic vesicle proteins, and the trans-Golgi network residual proteins in living axons at high resolution using laser scan microscopy of green fluorescent protein-tagged proteins after photobleaching.We found that all of these proteins are transported by tubulovesicular organelles of various sizes and shapes that circulate within axons from branch to branch and switch the direction of movement.These organelles are distinct from the endosomal compartments and constitute a new entity of membrane organelles that mediate the transport of newly synthesized proteins from the trans-Golgi network to the plasma membrane.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Anatomy, Graduate School of Medicine, University of Tokyo, Hongo, Tokyo, Japan, 113.

ABSTRACT
Newly synthesized membrane proteins are transported by fast axonal flow to their targets such as the plasma membrane and synaptic vesicles. However, their transporting vesicles have not yet been identified. We have successfully visualized the transporting vesicles of plasma membrane proteins, synaptic vesicle proteins, and the trans-Golgi network residual proteins in living axons at high resolution using laser scan microscopy of green fluorescent protein-tagged proteins after photobleaching. We found that all of these proteins are transported by tubulovesicular organelles of various sizes and shapes that circulate within axons from branch to branch and switch the direction of movement. These organelles are distinct from the endosomal compartments and constitute a new entity of membrane organelles that mediate the transport of newly synthesized proteins from the trans-Golgi network to the plasma membrane.

Show MeSH
Related in: MedlinePlus