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CAPS and syntaxin dock dense core vesicles to the plasma membrane in neurons.

Hammarlund M, Watanabe S, Schuske K, Jorgensen EM - J. Cell Biol. (2008)

Bottom Line: In Caenorhabditis elegans motor neurons, dense core vesicles dock at the plasma membrane but are excluded from active zones at synapses.Both the CAPS and UNC-13 docking pathways converge on syntaxin, a component of the SNARE (soluble N-ethyl-maleimide-sensitive fusion protein attachment receptor) complex.CAPS function in dense core vesicle docking parallels UNC-13 in synaptic vesicle docking, which suggests that these related proteins act similarly to promote docking of independent vesicle populations.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Utah, Salt Lake City, UT 84112, USA.

ABSTRACT
Docking to the plasma membrane prepares vesicles for rapid release. Here, we describe a mechanism for dense core vesicle docking in neurons. In Caenorhabditis elegans motor neurons, dense core vesicles dock at the plasma membrane but are excluded from active zones at synapses. We have found that the calcium-activated protein for secretion (CAPS) protein is required for dense core vesicle docking but not synaptic vesicle docking. In contrast, we see that UNC-13, a docking factor for synaptic vesicles, is not essential for dense core vesicle docking. Both the CAPS and UNC-13 docking pathways converge on syntaxin, a component of the SNARE (soluble N-ethyl-maleimide-sensitive fusion protein attachment receptor) complex. Overexpression of open syntaxin can bypass the requirement for CAPS in dense core vesicle docking. Thus, CAPS likely promotes the open state of syntaxin, which then docks dense core vesicles. CAPS function in dense core vesicle docking parallels UNC-13 in synaptic vesicle docking, which suggests that these related proteins act similarly to promote docking of independent vesicle populations.

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CAPS/UNC-31 functions with syntaxin to dock dense core vesicles. (A) UNC-31 is required for docking of dense core vesicles. (B) UNC-31 is not required for docking synaptic vesicles at active zones. (C) Syntaxin is required for docking dense core vesicles. In the mosaic strain, syntaxin is present in acetylcholine neurons but absent in GABA neurons. (D) Open syntaxin bypasses the requirement for UNC-31. DCV, dense core vesicle. (E) Dense core vesicles close to the plasma membrane but not docked (arrowheads) in unc-31. (F) A dense core vesicle that is docked (arrowhead) in unc-31 open syntaxin. Asterisks indicate approximate p-values (***, P < 0.001; **, P between 0.01 and 0.001; *, P between 0.05 and 0.01; see text for exact p-values). Error bars indicate SEM of the genotype mean. Syx, syntaxin; wt, wild type. Bars, 100 nm.
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fig4: CAPS/UNC-31 functions with syntaxin to dock dense core vesicles. (A) UNC-31 is required for docking of dense core vesicles. (B) UNC-31 is not required for docking synaptic vesicles at active zones. (C) Syntaxin is required for docking dense core vesicles. In the mosaic strain, syntaxin is present in acetylcholine neurons but absent in GABA neurons. (D) Open syntaxin bypasses the requirement for UNC-31. DCV, dense core vesicle. (E) Dense core vesicles close to the plasma membrane but not docked (arrowheads) in unc-31. (F) A dense core vesicle that is docked (arrowhead) in unc-31 open syntaxin. Asterisks indicate approximate p-values (***, P < 0.001; **, P between 0.01 and 0.001; *, P between 0.05 and 0.01; see text for exact p-values). Error bars indicate SEM of the genotype mean. Syx, syntaxin; wt, wild type. Bars, 100 nm.

Mentions: How do dense core vesicles dock to the plasma membrane? CAPS/UNC-31 is composed of a MUN domain, a pleckstrin homology phosphoinositide-binding domain, and a C2 phosphoinositide and calcium-binding motif (Ann et al., 1997; Speese et al., 2007). CAPS primes secretory granules in PC12 cells for calcium-stimulated release (Walent et al., 1992; Ann et al., 1997; Grishanin et al., 2004). In C. elegans, CAPS is essential for neuropeptide secretion but not synaptic vesicle secretion (Speese et al., 2007). It is possible that the defect in neuropeptide secretion in CAPS mutants is caused by a defect in docking of dense core vesicles. The mutation unc-31(e928) completely deletes the coding region for the CAPS protein (Charlie et al., 2006; Speese et al., 2007). We found that in the absence of CAPS, dense core vesicle docking is nearly eliminated (Fig. 4 A; docked dense core vesicles per profile: wild type, 0.057; unc-31(e928), 0.005; P < 0.0001). Lack of docking is not caused by a reduction in the total number of dense core vesicles because unc-31(e928) mutant animals have more total dense core vesicles than the wild type (Fig. 5 B; total dense core vesicles per profile: wild type, 0.95; unc-31(e928), 2.01; P < 0.0001). To confirm that the defect in unc-31(e928) in dense core vesicle docking is caused by a loss of UNC-31 rather than a potential background mutation, we performed a similar analysis on a second allele, unc-31(u280) (Fig. 4 A; Speese et al., 2007). No docked dense core vesicles were observed in the unc-31(u280) allele (docked dense core vesicles per profile: wild type, 0.057; unc-31(u280), 0.0; P < 0.0001). Furthermore, like unc-31(e928), unc-31(u280) accumulated total dense core vesicles (Fig. 5 C; total dense core vesicles per profile: wild type, 0.95; unc-31(u280), 1.19; P = 0.0006). The accumulation of dense core vesicles in unc-31 mutants is consistent with the proposed function of CAPS in dense core vesicle release. Similarly, Drosophila melanogaster CAPS mutants have an increased number of dense core vesicles in the boutons of neuromuscular junctions (Renden et al., 2001). The near-complete lack of docked dense core vesicles in the two unc-31 mutants demonstrates that CAPS is required for dense core vesicle docking.


CAPS and syntaxin dock dense core vesicles to the plasma membrane in neurons.

Hammarlund M, Watanabe S, Schuske K, Jorgensen EM - J. Cell Biol. (2008)

CAPS/UNC-31 functions with syntaxin to dock dense core vesicles. (A) UNC-31 is required for docking of dense core vesicles. (B) UNC-31 is not required for docking synaptic vesicles at active zones. (C) Syntaxin is required for docking dense core vesicles. In the mosaic strain, syntaxin is present in acetylcholine neurons but absent in GABA neurons. (D) Open syntaxin bypasses the requirement for UNC-31. DCV, dense core vesicle. (E) Dense core vesicles close to the plasma membrane but not docked (arrowheads) in unc-31. (F) A dense core vesicle that is docked (arrowhead) in unc-31 open syntaxin. Asterisks indicate approximate p-values (***, P < 0.001; **, P between 0.01 and 0.001; *, P between 0.05 and 0.01; see text for exact p-values). Error bars indicate SEM of the genotype mean. Syx, syntaxin; wt, wild type. Bars, 100 nm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2234227&req=5

fig4: CAPS/UNC-31 functions with syntaxin to dock dense core vesicles. (A) UNC-31 is required for docking of dense core vesicles. (B) UNC-31 is not required for docking synaptic vesicles at active zones. (C) Syntaxin is required for docking dense core vesicles. In the mosaic strain, syntaxin is present in acetylcholine neurons but absent in GABA neurons. (D) Open syntaxin bypasses the requirement for UNC-31. DCV, dense core vesicle. (E) Dense core vesicles close to the plasma membrane but not docked (arrowheads) in unc-31. (F) A dense core vesicle that is docked (arrowhead) in unc-31 open syntaxin. Asterisks indicate approximate p-values (***, P < 0.001; **, P between 0.01 and 0.001; *, P between 0.05 and 0.01; see text for exact p-values). Error bars indicate SEM of the genotype mean. Syx, syntaxin; wt, wild type. Bars, 100 nm.
Mentions: How do dense core vesicles dock to the plasma membrane? CAPS/UNC-31 is composed of a MUN domain, a pleckstrin homology phosphoinositide-binding domain, and a C2 phosphoinositide and calcium-binding motif (Ann et al., 1997; Speese et al., 2007). CAPS primes secretory granules in PC12 cells for calcium-stimulated release (Walent et al., 1992; Ann et al., 1997; Grishanin et al., 2004). In C. elegans, CAPS is essential for neuropeptide secretion but not synaptic vesicle secretion (Speese et al., 2007). It is possible that the defect in neuropeptide secretion in CAPS mutants is caused by a defect in docking of dense core vesicles. The mutation unc-31(e928) completely deletes the coding region for the CAPS protein (Charlie et al., 2006; Speese et al., 2007). We found that in the absence of CAPS, dense core vesicle docking is nearly eliminated (Fig. 4 A; docked dense core vesicles per profile: wild type, 0.057; unc-31(e928), 0.005; P < 0.0001). Lack of docking is not caused by a reduction in the total number of dense core vesicles because unc-31(e928) mutant animals have more total dense core vesicles than the wild type (Fig. 5 B; total dense core vesicles per profile: wild type, 0.95; unc-31(e928), 2.01; P < 0.0001). To confirm that the defect in unc-31(e928) in dense core vesicle docking is caused by a loss of UNC-31 rather than a potential background mutation, we performed a similar analysis on a second allele, unc-31(u280) (Fig. 4 A; Speese et al., 2007). No docked dense core vesicles were observed in the unc-31(u280) allele (docked dense core vesicles per profile: wild type, 0.057; unc-31(u280), 0.0; P < 0.0001). Furthermore, like unc-31(e928), unc-31(u280) accumulated total dense core vesicles (Fig. 5 C; total dense core vesicles per profile: wild type, 0.95; unc-31(u280), 1.19; P = 0.0006). The accumulation of dense core vesicles in unc-31 mutants is consistent with the proposed function of CAPS in dense core vesicle release. Similarly, Drosophila melanogaster CAPS mutants have an increased number of dense core vesicles in the boutons of neuromuscular junctions (Renden et al., 2001). The near-complete lack of docked dense core vesicles in the two unc-31 mutants demonstrates that CAPS is required for dense core vesicle docking.

Bottom Line: In Caenorhabditis elegans motor neurons, dense core vesicles dock at the plasma membrane but are excluded from active zones at synapses.Both the CAPS and UNC-13 docking pathways converge on syntaxin, a component of the SNARE (soluble N-ethyl-maleimide-sensitive fusion protein attachment receptor) complex.CAPS function in dense core vesicle docking parallels UNC-13 in synaptic vesicle docking, which suggests that these related proteins act similarly to promote docking of independent vesicle populations.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Utah, Salt Lake City, UT 84112, USA.

ABSTRACT
Docking to the plasma membrane prepares vesicles for rapid release. Here, we describe a mechanism for dense core vesicle docking in neurons. In Caenorhabditis elegans motor neurons, dense core vesicles dock at the plasma membrane but are excluded from active zones at synapses. We have found that the calcium-activated protein for secretion (CAPS) protein is required for dense core vesicle docking but not synaptic vesicle docking. In contrast, we see that UNC-13, a docking factor for synaptic vesicles, is not essential for dense core vesicle docking. Both the CAPS and UNC-13 docking pathways converge on syntaxin, a component of the SNARE (soluble N-ethyl-maleimide-sensitive fusion protein attachment receptor) complex. Overexpression of open syntaxin can bypass the requirement for CAPS in dense core vesicle docking. Thus, CAPS likely promotes the open state of syntaxin, which then docks dense core vesicles. CAPS function in dense core vesicle docking parallels UNC-13 in synaptic vesicle docking, which suggests that these related proteins act similarly to promote docking of independent vesicle populations.

Show MeSH
Related in: MedlinePlus