7 research outputs found
The yeast endocytic early/sorting compartment exists as an independent sub-compartment within the trans-Golgi network
Although budding yeast has been extensively used as a model organism for studying organelle functions and intracellular vesicle trafficking, whether it possesses an independent endocytic early/sorting compartment that sorts endocytic cargos to the endo-lysosomal pathway or the recycling pathway has long been unclear. The structure and properties of the endocytic early/sorting compartment differ significantly between organisms; in plant cells, the trans-Golgi network (TGN) serves this role, whereas in mammalian cells a separate intracellular structure performs this function. The yeast syntaxin homolog Tlg2p, widely localizing to the TGN and endosomal compartments, is presumed to act as a Q-SNARE for endocytic vesicles, but which compartment is the direct target for endocytic vesicles remained unanswered. Here we demonstrate by high-speed and high-resolution 4D imaging of fluorescently labeled endocytic cargos that the Tlg2p-residing compartment within the TGN functions as the early/sorting compartment. After arriving here, endocytic cargos are recycled to the plasma membrane or transported to the yeast Rab5-residing endosomal compartment through the pathway requiring the clathrin adaptors GGAs. Interestingly, Gga2p predominantly localizes at the Tlg2p-residing compartment, and the deletion of GGAs has little effect on another TGN region where Sec7p is present but suppresses dynamics of the Tlg2-residing early/sorting compartment, indicating that the Tlg2p- and Sec7p-residing regions are discrete entities in the mutant. Thus, the Tlg2p-residing region seems to serve as an early/sorting compartment and function independently of the Sec7p-residing region within the TGN
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The yeast endocytic early/sorting compartment exists as an independent sub-compartment within the trans-Golgi network
Although budding yeast has been extensively used as a model organism for studying organelle functions and intracellular vesicle trafficking, whether it possesses an independent endocytic early/sorting compartment that sorts endocytic cargos to the endo-lysosomal pathway or the recycling pathway has long been unclear. The structure and properties of the endocytic early/sorting compartment differ significantly between organisms; in plant cells the trans -Golgi network (TGN) serves this role, whereas in mammalian cells a separate intracellular structure performs this function. The yeast syntaxin homolog Tlg2p, widely localizing to the TGN and endosomal compartments, is presumed to act as a Q-SNARE for endocytic vesicles, but which compartment is the direct target for endocytic vesicles remained unanswered. Here we demonstrate by high-speed and high-resolution 4D imaging of fluorescently labeled endocytic cargos that the Tlg2p-residing compartment within the TGN functions as the early/sorting compartment. After arriving here, endocytic cargos are recycled to the plasma membrane or transported to the yeast Rab5-residing endosomal compartment through the pathway requiring the clathrin adaptors GGAs. Interestingly, Gga2p predominantly localizes at the Tlg2p-residing compartment, and the deletion of GGAs has little effect on another TGN region where Sec7p is present but suppresses dynamics of the Tlg2-residing early/sorting compartment, indicating that the Tlg2p- and Sec7p-residing regions are discrete entities in the mutant. Thus, the Tlg2p-residing region seems to serve as an early/sorting compartment, and function independently of the Sec7p-residing region within the TGN
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The yeast endocytic early/sorting compartment exists as an independent sub-compartment within the trans-Golgi network.
Although budding yeast has been extensively used as a model organism for studying organelle functions and intracellular vesicle trafficking, whether it possesses an independent endocytic early/sorting compartment that sorts endocytic cargos to the endo-lysosomal pathway or the recycling pathway has long been unclear. The structure and properties of the endocytic early/sorting compartment differ significantly between organisms; in plant cells, the trans-Golgi network (TGN) serves this role, whereas in mammalian cells a separate intracellular structure performs this function. The yeast syntaxin homolog Tlg2p, widely localizing to the TGN and endosomal compartments, is presumed to act as a Q-SNARE for endocytic vesicles, but which compartment is the direct target for endocytic vesicles remained unanswered. Here we demonstrate by high-speed and high-resolution 4D imaging of fluorescently labeled endocytic cargos that the Tlg2p-residing compartment within the TGN functions as the early/sorting compartment. After arriving here, endocytic cargos are recycled to the plasma membrane or transported to the yeast Rab5-residing endosomal compartment through the pathway requiring the clathrin adaptors GGAs. Interestingly, Gga2p predominantly localizes at the Tlg2p-residing compartment, and the deletion of GGAs has little effect on another TGN region where Sec7p is present but suppresses dynamics of the Tlg2-residing early/sorting compartment, indicating that the Tlg2p- and Sec7p-residing regions are discrete entities in the mutant. Thus, the Tlg2p-residing region seems to serve as an early/sorting compartment and function independently of the Sec7p-residing region within the TGN
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Paxillin phosphorylation counteracts proteoglycan-mediated inhibition of axon regeneration
In the adult central nervous system, the tips of axons severed by injury are commonly transformed into dystrophic endballs and cease migration upon encountering a rising concentration gradient of inhibitory proteoglycans. However, intracellular signaling networks mediating endball migration failure remain largely unknown. Here we show that manipulation of protein kinase A (PKA) or its downstream adhesion component paxillin can reactivate the locomotive machinery of endballs in vitro and facilitate axon growth after injury in vivo. In dissociated cultures of adult rat dorsal root ganglion neurons, PKA is activated in endballs formed on gradients of the inhibitory proteoglycan aggrecan, and pharmacological inhibition of PKA promotes axon growth on aggrecan gradients most likely through phosphorylation of paxillin at serine 301. Remarkably, pre-formed endballs on aggrecan gradients resume forward migration in response to PKA inhibition. This resumption of endball migration is associated with increased turnover of adhesive point contacts dependent upon paxillin phosphorylation. Furthermore, expression of phosphomimetic paxillin overcomes aggrecan-mediated growth arrest of endballs, and facilitates axon growth after optic nerve crush in vivo. These results point to the importance of adhesion dynamics in restoring endball migration and suggest a potential therapeutic target for axon tract repair.
•Inhibitory proteoglycan gradients cause PKA activation in dystrophic endballs.•PKA inhibition restores the growth capability of dystrophic endballs.•Endballs resume forward migration possibly through increased adhesion dynamics.•Serine phosphorylation on paxillin mediates resumption of endball migration.•Paxillin phosphorylation promotes axon growth in the injured CNS