Synaptotagmin VII Regulates Ca2+-Dependent Exocytosis of Lysosomes in Fibroblasts

Martínez, Iñigo; Chakrabarti, Sabyasachi; Hellevik, Turid; Morehead, Jennifer; Fowler, Kimberly T.; Andrews, Norma W.

doi:10.1083/jcb.148.6.1141

Cited by 281 publications

(349 citation statements)

References 37 publications

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“…This was confirmed by the weak co-localization found between several FM dyes and fluorescent endocytic markers (Fig. 1B) (20). Five-minute co-application of green FM1-43 and 3,000-MW Texas Red dextran (Tx-d) resulted in a lower correlation coefficient (r 12 ϭ 0.16 Ϯ 0.07, n ϭ 8; see SI Appendix) than that found when coapplying green FM1-43 and red FM4-64 ( Fig.…”

Section: Resultssupporting

confidence: 53%

FM dyes enter via a store-operated calcium channel and modify calcium signaling of cultured astrocytes

Hérault

Oheim

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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The amphiphilic fluorescent styryl pyridinium dyes FM1-43 and FM4-64 are used to probe activity-dependent synaptic vesicle cycling in neurons. Cultured astrocytes can internalize FM1-43 and FM4-64 inside vesicles but their uptake is insensitive to the elevation of cytosolic calcium (Ca 2؉ ) concentration and the underlying mechanism remains unclear. Here we used total internal reflection fluorescence microscopy and pharmacological tools to study the mechanisms of FM4-64 uptake into cultured astrocytes from mouse neocortex. Our data show that: (i) endocytosis is not a major route for FM4-64 uptake into astrocytes; (ii) FM4-64 enters astrocytes through an aqueous pore and strongly affects Ca 2؉ homeostasis; (iii) partitioning of FM4-64 into the outer leaflet of the plasma membrane results in a facilitation of store-operated Ca 2؉ entry (SOCE) channel gating; (iv) FM4-64 permeates and competes with Ca 2؉ for entry through a SOCE channel; (v) intracellular FM4-64 mobilizes Ca 2؉ from the endoplasmic reticulum stores, conveying a positive feedback to activate SOCE and to sustain dye uptake into astrocytes. Our study demonstrates that FM dyes are not markers of cycling vesicles in astrocytes and calls for a careful interpretation of FM fluorescence.calcium homeostasis ͉ endoplasmic reticulum ͉ lipid bilayer ͉ styryl dye ͉ TIRF I n astrocytes the strong expression of the plasma membrane (PM) store-operated Ca 2ϩ entry (SOCE) (1-4) contrasts with a relatively small expression of voltage-gated channels and AMPA/ NMDA ligand-gated channels that are mostly expressed by neurons. In response to neuronal activity and neurotransmitter release, the activation of astroglial metabotropic receptors induces a reduction of the endoplasmic reticulum (ER) Ca 2ϩ concentration ([Ca 2ϩ ] ER ) that leads to the activation of the SOCE and to an elevation of the intracellular Ca 2ϩ concentration ([Ca 2ϩ ] i ) (4) that is critical for the electrically silent astrocytes to release gliotransmitters and control nearby neurons, glia, and blood vessels (5). The type 1-2 stromal interaction molecules (STIM1-2) and Orai1-3 membrane proteins are key components of the SOCE, being the ER Ca 2ϩ sensor and the PM pore-forming subunit, respectively (6, 7). A possible participation of the classical transient receptor potential (TRPC) channels to the protein complex generating the SOCE is debated in several cell types (8), including astrocytes (3).Styryl pyridinium FM dyes are amphiphilic molecules that reversibly partition in lipid membranes. Their positively charged pyridinium head prevents them from diffusing through the PM. Only weakly fluorescent in aqueous solution their quantum yield increases in a lipid environment. Their activity-dependent uptake and destaining has made them common probes for synaptic vesicle cycling at the nerve terminals (9-11). Cultured astrocytes internalize FM dyes (3,(12)(13)(14)(15)(16)(17)(18). However, this uptake is not modified by the activation of metabotropic glutamate receptors (19), and its mechanism remains elusiv...

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Section: Resultssupporting

confidence: 53%

FM dyes enter via a store-operated calcium channel and modify calcium signaling of cultured astrocytes

Hérault

Oheim

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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“…Regulated exocytosis of granules with lysosomal characteristics has been more frequently described in cells of the hematopoietic lineage, such as platelets, mast cells, neutrophils, and cytotoxic lymphocytes (22). Recent evidence indicates, however, that intracellular free Ca 2ϩ elevations can trigger exocytosis of conventional lysosomes in many cell types and that the process may be regulated by a transmembrane protein with Ca 2ϩ -binding properties, synaptotagmin VII (19,26). Taken together, the available evidence suggests that activation of host cell signaling pathways and Ca 2ϩ elevation at the site of parasite attachment results in localized lysosome recruitment, docking, and fusion, events that contribute to formation of the T. cruzi-containing intracellular vacuole.…”

Section: Discussionmentioning

confidence: 99%

Dual Role of Signaling Pathways Leading to Ca²⁺and Cyclic AMP Elevation in Host Cell Invasion byTrypanosoma cruzi

et al. 2000

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Cell invasion by the protozoan parasite Trypanosoma cruzi involves activation of host signaling pathways and the recruitment and fusion of lysosomes at the parasite entry site. A major signaling pathway regulating invasion of fibroblasts, epithelial cells, and myoblasts involves mobilization of Ca 2؉ from intracellular stores and requires the activity of a T. cruzi serine peptidase, oligopeptidase B (OPB). Deletion of the OPB gene results in a marked defect in trypomastigote virulence, consistent with a greatly reduced cell invasion capacity. Here we show that uptake by macrophages, on the other hand, is largely independent of OPB expression and sensitive to inhibition of by cytochalasin D. The residual invasion capacity of OPBnull trypomastigotes in fibroblasts still involves lysosome recruitment, although in a significantly delayed fashion. Transient elevations in intracellular Ca 2؉ concentrations were observed in host cells exposed to both wild-type and OPBnull trypomastigotes, but the signals triggered by the mutant parasites were less vigorous and delayed. The capacity of triggering elevation in host cell cyclic AMP (cAMP), however, was unaltered in OPBnull trypomastigotes. Modulation in cAMP levels preferentially affected the residual cell invasion capacity of OPBnull parasites, suggesting that this signaling pathway can play a dominant role in promoting cell invasion in the absence of the major OPB-dependent pathway.

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“…Synaptotagmin VII is known to regulate lysosomal exocytosis; 29,38 however, Syt VII expression in astrocytes is disputed. 27,28 Multiple Syt VII isoforms reported in neurons and chromaffin cells.…”

Section: Kda-tritc Dextran 500kda-fitc Dextranmentioning

confidence: 99%

Injured astrocytes are repaired by Synaptotagmin XI-regulated lysosome exocytosis

et al. 2015

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Astrocytes are known to facilitate repair following brain injury; however, little is known about how injured astrocytes repair themselves. Repair of cell membrane injury requires Ca 2+ -triggered vesicle exocytosis. In astrocytes, lysosomes are the main Ca 2+ -regulated exocytic vesicles. Here we show that astrocyte cell membrane injury results in a large and rapid calcium increase. This triggers robust lysosome exocytosis where the fusing lysosomes release all luminal contents and merge fully with the plasma membrane. In contrast to this, receptor stimulation produces a small sustained calcium increase, which is associated with partial release of the lysosomal luminal content, and the lysosome membrane does not merge into the plasma membrane. In most cells, lysosomes express the synaptotagmin (Syt) isoform Syt VII; however, this isoform is not present on astrocyte lysosomes and exogenous expression of Syt VII on lysosome inhibits their exocytosis. Deletion of one of the most abundant Syt isoform in astrocyte -Syt XI -suppresses astrocyte lysosome exocytosis. This identifies lysosome as Syt XI-regulated exocytic vesicle in astrocytes. Further, inhibition of lysosome exocytosis (by Syt XI depletion or Syt VII expression) prevents repair of injured astrocytes. These results identify the lysosomes and Syt XI as the sub-cellular and molecular regulators, respectively of astrocyte cell membrane repair.

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Synaptotagmin VII Regulates Ca2+-Dependent Exocytosis of Lysosomes in Fibroblasts

Cited by 281 publications

References 37 publications

FM dyes enter via a store-operated calcium channel and modify calcium signaling of cultured astrocytes

FM dyes enter via a store-operated calcium channel and modify calcium signaling of cultured astrocytes

Dual Role of Signaling Pathways Leading to Ca²⁺and Cyclic AMP Elevation in Host Cell Invasion byTrypanosoma cruzi

Injured astrocytes are repaired by Synaptotagmin XI-regulated lysosome exocytosis

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Synaptotagmin VII Regulates Ca2+-Dependent Exocytosis of Lysosomes in Fibroblasts

Cited by 281 publications

References 37 publications

FM dyes enter via a store-operated calcium channel and modify calcium signaling of cultured astrocytes

FM dyes enter via a store-operated calcium channel and modify calcium signaling of cultured astrocytes

Dual Role of Signaling Pathways Leading to Ca2+and Cyclic AMP Elevation in Host Cell Invasion byTrypanosoma cruzi

Injured astrocytes are repaired by Synaptotagmin XI-regulated lysosome exocytosis

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Dual Role of Signaling Pathways Leading to Ca²⁺and Cyclic AMP Elevation in Host Cell Invasion byTrypanosoma cruzi