The <scp>SNARE</scp> protein vti1a functions in dense‐core vesicle biogenesis

Walter, Alexander; Kurps, Julia; Wit, Heidi de; Schöning, Susanne; Toft-Bertelsen, Trine L.; Lauks, Juliane; Ziomkiewicz, Iwona; Weiss, Annita Ngatchou; Schulz, Alexander; Mollard, Gabriele Fischer von; Verhage, Matthijs; Sørensen, Jakob B.

doi:10.15252/embj.201387549

Cited by 36 publications

(49 citation statements)

References 64 publications

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“…3B). Consistent with this, we quantified the intensities of CgA in cytosolic individual granules that are defined as LDCVs (Walter et al, 2014) and showed that the CgA intensity in the LDCV proportion was increased in muted-KD PC12 cells compared with the control cells (Fig. 3C).…”

Section: Morphological Changes Of Ldcvs In Mu Chromaffin Cellssupporting

confidence: 72%

Impaired maturation of large dense-core vesicles in muted-deficient adrenal chromaffin cells

Hao

Wei

Feng

et al. 2015

Journal of Cell Science

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The large dense-core vesicle (LDCV), a type of lysosome-related organelle, is involved in the secretion of hormones and neuropeptides in specialized secretory cells. The granin family is a driving force in LDCV biogenesis, but the machinery for granin sorting to this biogenesis pathway is largely unknown. The mu mutant mouse, which carries a spontaneous null mutation on the Muted gene (also known as Bloc1s5), which encodes a subunit of the biogenesis of lysosome-related organelles complex-1 (BLOC-1), is a mouse model of Hermansky-Pudlak syndrome. Here, we found that LDCVs were enlarged in mu adrenal chromaffin cells. Chromogranin A (CgA, also known as CHGA) was increased in mu adrenals and mutedknockdown cells. The increased CgA in mu mice was likely due a failure to export this molecule out of immature LDCVs, which impairs LDCV maturation and docking. In mu chromaffin cells, the size of readily releasable pool and the vesicle release frequency were reduced. Our studies suggest that the muted protein is involved in the selective export of CgA during the biogenesis of LDCVs.

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Section: Morphological Changes Of Ldcvs In Mu Chromaffin Cellssupporting

confidence: 72%

Impaired maturation of large dense-core vesicles in muted-deficient adrenal chromaffin cells

Hao

Wei

Feng

et al. 2015

Journal of Cell Science

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“…Immature DCVs bud off from the trans‐Golgi and undergo several maturation steps, including homotypic fusion with other immature DCVs, vesicle acidification, peptide processing and sorting of cargos . In recent years, a small but growing number of proteins have been identified as playing roles in DCV biogenesis, beginning to shed some light on the molecular mechanisms underlying the steps in DCV maturation . In one approach to identifying proteins important for DCV biogenesis, genetic screens were performed in Caenorhabditis elegans .…”

Section: Introductionmentioning

confidence: 99%

“…[1][2][3][4] In recent years, a small but growing number of proteins have been identified as playing roles in DCV biogenesis, beginning to shed some light on the molecular mechanisms underlying the steps in DCV maturation. [5][6][7][8][9][10][11][12][13][14][15][16][17][18] In one approach to identifying proteins important for DCV biogenesis, genetic screens were performed in Caenorhabditis elegans. [19][20][21][22][23][24][25] These screens identified several molecules important for early stages of DCV biogenesis and maturation, including the small G-protein RAB-2 and several RAB-2 effectors, including the conserved coiled-coil protein CCCP-1.…”

Section: Introductionmentioning

confidence: 99%

The dense‐core vesicle maturation protein CCCP‐1 binds RAB‐2 and membranes through its C‐terminal domain

Cattin‐Ortolá

Topalidou

Dosey

et al. 2017

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Dense-core vesicles (DCVs) are secretory organelles that store and release modulatory neurotransmitters from neurons and endocrine cells. Recently, the conserved coiled-coil protein CCCP-1 was identified as a component of the DCV biogenesis pathway in the nematode C. elegans. CCCP-1 binds the small GTPase RAB-2 and colocalizes with it at the trans-Golgi. Here we report a structure-function analysis of CCCP-1 to identify domains of the protein important for its localization, binding to RAB-2, and function in DCV biogenesis. We find that the CCCP-1 C-terminal domain (CC3) has multiple activities. CC3 is necessary and sufficient for CCCP-1 localization and for binding to RAB-2, and is required for the function of CCCP-1 in DCV biogenesis. Additionally, CCCP-1 binds membranes directly through its CC3 domain, indicating that CC3 may comprise a previously uncharacterized lipid-binding motif. We conclude that CCCP-1 is a coiled-coil protein that binds an activated Rab and localizes to the Golgi via its C-terminus, properties similar to members of the golgin family of proteins. CCCP-1 also shares biophysical features with golgins; it has an elongated shape and forms oligomers.

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“…3d, e, b, f, h, and k). Vti1a is a SNARE that is localized in the TGN and is involved in vesicle generation during exocytosis and the retrograde transport from early and late endosomes to the TGN (Malsam and Sollner, 2011;Walter et al, 2014). Vti1b, another SNARE, is localized in the TGN and late endosomes, and participates in post-Golgi secretory trafficking and fusion of late endosomes (Antonin et al, 2000;Murray et al, 2005).…”

Section: Isolation Of the Fc-fusion Protein From Intracellular Fractionsmentioning

confidence: 99%

Site‐specific monitoring of N‐Glycosylation profiles of a CTLA4‐Fc‐fusion protein from the secretory pathway to the extracellular environment

Oliveira¹,

Spencer²,

Barton³

et al. 2017

Biotech & Bioengineering

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Glycosylation often plays a key role in the safety and efficacy of therapeutic proteins to patients, thus underlying the need for consistent control of this important post-translational modification during biologics production. In this study, we profiled the site-specific evolution of N-glycans on a CTLA4-Fc-fusion protein, from the intracellular secretory pathway to the conditioned medium (CM) in fed-batch cell culture. For this, we developed an approach that combined sub-cellular fractionation with liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses. The study revealed that there was a significant amount of heterogeneity in the glycans displayed amongst the three distinct N-glycosylation sites. Furthermore, 54-60% of the intracellular protein was characterized by Man8 and Man9 glycans on day 10, when the cell density peaks, indicative of a significant bottleneck between the endoplasmic reticulum (ER) and the cis-Golgi. At longer culture duration, the accumulation of intracellular protein with bi-antennary-fucosylated GlcNAc-terminated residues identified the formation of another bottleneck in the medial and trans-Golgi compartments, which subsequently led to a decrease in sialylated species in the secreted protein. Glucose deprivation caused a reduction in the Man8 and Man9 glycans in favor of Man5 glycans and bi-antennary-fucosylated GlcNAc-terminated residues in the organellar pool of the Fc-fusion protein. However, transient deprivation of glucose did not lead to major differences in the glycan profile of proteins secreted into the CM. The approach developed here allows us to probe the secretory pathway and sheds light on the site-specific intracellular processing of glycans during fed-batch cell culture, thus serving as an initial step towards their rational control. Biotechnol. Bioeng. 2017;114: 1550-1560. © 2017 Wiley Periodicals, Inc.

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The SNARE protein vti1a functions in dense‐core vesicle biogenesis

Cited by 36 publications

References 64 publications

Impaired maturation of large dense-core vesicles in muted-deficient adrenal chromaffin cells

Impaired maturation of large dense-core vesicles in muted-deficient adrenal chromaffin cells

The dense‐core vesicle maturation protein CCCP‐1 binds RAB‐2 and membranes through its C‐terminal domain

Site‐specific monitoring of N‐Glycosylation profiles of a CTLA4‐Fc‐fusion protein from the secretory pathway to the extracellular environment

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