A kinetic view of clathrin assembly and endocytic cargo sorting

2021 ◽  
Vol 71 ◽  
pp. 130-138
Author(s):  
Min Wu ◽  
Xudong Wu
Keyword(s):  
Cargo Sorting

scholarly journals Cargo sorting zones in the trans-Golgi network visualized by super-resolution confocal live imaging microscopy in plants

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yutaro Shimizu ◽  
Junpei Takagi ◽  
Emi Ito ◽  
Yoko Ito ◽  
Kazuo Ebine ◽  
...  
Keyword(s):  
Membrane Trafficking ◽  
High Speed ◽  
Super Resolution ◽  
Adaptor Protein ◽  
Transport Proteins ◽  
3D Localization ◽  
Golgi Network ◽  
Distinct Distribution ◽  
Vacuolar Trafficking ◽  
Cargo Sorting

AbstractThe trans-Golgi network (TGN) has been known as a key platform to sort and transport proteins to their final destinations in post-Golgi membrane trafficking. However, how the TGN sorts proteins with different destinies still remains elusive. Here, we examined 3D localization and 4D dynamics of TGN-localized proteins of Arabidopsis thaliana that are involved in either secretory or vacuolar trafficking from the TGN, by a multicolor high-speed and high-resolution spinning-disk confocal microscopy approach that we developed. We demonstrate that TGN-localized proteins exhibit spatially and temporally distinct distribution. VAMP721 (R-SNARE), AP (adaptor protein complex)−1, and clathrin which are involved in secretory trafficking compose an exclusive subregion, whereas VAMP727 (R-SNARE) and AP-4 involved in vacuolar trafficking compose another subregion on the same TGN. Based on these findings, we propose that the single TGN has at least two subregions, or “zones”, responsible for distinct cargo sorting: the secretory-trafficking zone and the vacuolar-trafficking zone.

scholarly journals Cofilin recruits F-actin to SPCA1 and promotes Ca2+-mediated secretory cargo sorting

2014 ◽  
Vol 206 (5) ◽  
pp. 635-654 ◽  
Author(s):  
Christine Kienzle ◽  
Nirakar Basnet ◽  
Alvaro H. Crevenna ◽  
Gisela Beck ◽  
Bianca Habermann ◽  
...  
Keyword(s):  
Secretory Pathway ◽  
Nitrilotriacetic Acid ◽  
Calcium Atpase ◽  
Secretory Proteins ◽  
Dependent Manner ◽  
Agarose Beads ◽  
P Type ◽  
Golgi Network ◽  
Phosphorylation Domain ◽  
Cargo Sorting

The actin filament severing protein cofilin-1 (CFL-1) is required for actin and P-type ATPase secretory pathway calcium ATPase (SPCA)-dependent sorting of secretory proteins at the trans-Golgi network (TGN). How these proteins interact and activate the pump to facilitate cargo sorting, however, is not known. We used purified proteins to assess interaction of the cytoplasmic domains of SPCA1 with actin and CFL-1. A 132–amino acid portion of the SPCA1 phosphorylation domain (P-domain) interacted with actin in a CFL-1–dependent manner. This domain, coupled to nickel nitrilotriacetic acid (Ni-NTA) agarose beads, specifically recruited F-actin in the presence of CFL-1 and, when expressed in HeLa cells, inhibited Ca2+ entry into the TGN and secretory cargo sorting. Mutagenesis of four amino acids in SPCA1 that represent the CFL-1 binding site also affected Ca2+ import into the TGN and secretory cargo sorting. Altogether, our findings reveal the mechanism of CFL-1–dependent recruitment of actin to SPCA1 and the significance of this interaction for Ca2+ influx and secretory cargo sorting.

scholarly journals pH-dependent Cargo Sorting from the Golgi

2011 ◽  
Vol 286 (12) ◽  
pp. 10058-10065 ◽  
Author(s):  
Chunjuan Huang ◽  
Amy Chang
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Atpase Activity ◽  
Secretory Pathway ◽  
Ph Dependence ◽  
Glucose Deprivation ◽  
Ph Homeostasis ◽  
Plasma Membrane Proteins ◽  
Cytosolic Ph ◽  
Cargo Sorting

The vacuolar proton-translocating ATPase (V-ATPase) plays a major role in organelle acidification and works together with other ion transporters to maintain pH homeostasis in eukaryotic cells. We analyzed a requirement for V-ATPase activity in protein trafficking in the yeast secretory pathway. Deficiency of V-ATPase activity caused by subunit deletion or glucose deprivation results in missorting of newly synthesized plasma membrane proteins Pma1 and Can1 directly from the Golgi to the vacuole. Vacuolar mislocalization of Pma1 is dependent on Gga adaptors although no Pma1 ubiquitination was detected. Proper cell surface targeting of Pma1 was rescued in V-ATPase-deficient cells by increasing the pH of the medium, suggesting that missorting is the result of aberrant cytosolic pH. In addition to mislocalization of the plasma membrane proteins, Golgi membrane proteins Kex2 and Vrg4 are also missorted to the vacuole upon loss of V-ATPase activity. Because the missorted cargos have distinct trafficking routes, we suggest a pH dependence for multiple cargo sorting events at the Golgi.

scholarly journals HID-1 controls formation of large dense core vesicles by influencing cargo sorting and trans-Golgi network acidification

2017 ◽  
Vol 28 (26) ◽  
pp. 3870-3880 ◽  
Author(s):  
Blake H. Hummer ◽  
Noah F. de Leeuw ◽  
Christian Burns ◽  
Lan Chen ◽  
Matthew S. Joens ◽  
...  
Keyword(s):  
Biochemical Properties ◽  
Neuroendocrine Cells ◽  
Dense Core ◽  
Core Formation ◽  
Dense Core Vesicles ◽  
Atpase Subunit ◽  
Trans Golgi Network ◽  
Large Dense Core Vesicles ◽  
Golgi Network ◽  
Cargo Sorting

Large dense core vesicles (LDCVs) mediate the regulated release of neuropeptides and peptide hormones. They form at the trans-Golgi network (TGN), where their soluble content aggregates to form a dense core, but the mechanisms controlling biogenesis are still not completely understood. Recent studies have implicated the peripheral membrane protein HID-1 in neuropeptide sorting and insulin secretion. Using CRISPR/Cas9, we generated HID-1 KO rat neuroendocrine cells, and we show that the absence of HID-1 results in specific defects in peptide hormone and monoamine storage and regulated secretion. Loss of HID-1 causes a reduction in the number of LDCVs and affects their morphology and biochemical properties, due to impaired cargo sorting and dense core formation. HID-1 KO cells also exhibit defects in TGN acidification together with mislocalization of the Golgi-enriched vacuolar H+-ATPase subunit isoform a2. We propose that HID-1 influences early steps in LDCV formation by controlling dense core formation at the TGN.

scholarly journals A cargo-sorting DNA robot

Science ◽  
2017 ◽  
Vol 357 (6356) ◽  
pp. eaan6558 ◽  
Author(s):  
Anupama J. Thubagere ◽  
Wei Li ◽  
Robert F. Johnson ◽  
Zibo Chen ◽  
Shayan Doroudi ◽  
...  
Keyword(s):  
Cargo Sorting

scholarly journals The Endosome-Associated Protein Hrs Is Hexameric and Controls Cargo Sorting as a “Master Molecule”

Structure ◽  
2006 ◽  
Vol 14 (4) ◽  
pp. 661-671 ◽  
Author(s):  
Lee Pullan ◽  
Srinivas Mullapudi ◽  
Zhong Huang ◽  
Philip R. Baldwin ◽  
Christopher Chin ◽  
...  
Keyword(s):  
Cargo Sorting

scholarly journals ARFGAP1 promotes the formation of COPI vesicles, suggesting function as a component of the coat

2002 ◽  
Vol 159 (1) ◽  
pp. 69-78 ◽  
Author(s):  
Jia-Shu Yang ◽  
Stella Y. Lee ◽  
Minggeng Gao ◽  
Sylvain Bourgoin ◽  
Paul A. Randazzo ◽  
...  
Keyword(s):  
Coat Protein ◽  
Gap Functions ◽  
Vesicle Formation ◽  
Golgi Membrane ◽  
Gtpase Activating Protein ◽  
Cargo Proteins ◽  
Adp Ribosylation Factor ◽  
Soluble Components ◽  
Cargo Sorting

The role of GTPase-activating protein (GAP) that deactivates ADP-ribosylation factor 1 (ARF1) during the formation of coat protein I (COPI) vesicles has been unclear. GAP is originally thought to antagonize vesicle formation by triggering uncoating, but later studies suggest that GAP promotes cargo sorting, a process that occurs during vesicle formation. Recent models have attempted to reconcile these seemingly contradictory roles by suggesting that cargo proteins suppress GAP activity during vesicle formation, but whether GAP truly antagonizes coat recruitment in this process has not been assessed directly. We have reconstituted the formation of COPI vesicles by incubating Golgi membrane with purified soluble components, and find that ARFGAP1 in the presence of GTP promotes vesicle formation and cargo sorting. Moreover, the presence of GTPγS not only blocks vesicle uncoating but also vesicle formation by preventing the proper recruitment of GAP to nascent vesicles. Elucidating how GAP functions in vesicle formation, we find that the level of GAP on the reconstituted vesicles is at least as abundant as COPI and that GAP binds directly to the dilysine motif of cargo proteins. Collectively, these findings suggest that ARFGAP1 promotes vesicle formation by functioning as a component of the COPI coat.

scholarly journals Epstein-Barr virus LMP1 modulates the CD63 interactome

2021 ◽  
Author(s):  
Mujeeb Cheerathodi ◽  
Dingani Nkosi ◽  
Allaura S. Cone ◽  
Sara B. York ◽  
David G. Meckes Jr.
Keyword(s):  
Intracellular Signaling ◽  
Ribosomal Proteins ◽  
Protein Localization ◽  
Cell Communication ◽  
Enrichment Analysis ◽  
Surface Proteins ◽  
Rab Gtpases ◽  
Interacting Proteins ◽  
Sorting Nexins ◽  
Cargo Sorting

Abstract Tetraspanin CD63 is a cluster of cell surface proteins with four transmembrane domains which associates with tetraspanin-enriched microdomains and typically localizes to late endosomes and lysosomes. CD63 plays an important role in cellular trafficking of different proteins, EV cargo sorting and vesicles formation. We have preciously shown that CD63 is important in LMP1 trafficking to EVs and this also affects LMP1 mediated intracellular signaling including MAPK/ERK, NF-κB and mTOR activation. Using the BioID combined with mass spectrometry, we sought to define the broad CD63 interactome and how LMP1 modulates this network of interacting proteins. We identified a total of 1600 total proteins as proximal interacting newtwork of proteins to CD63. Biological process enrichment analysis revealed significant involvement in signal transduction, cell communication, protein metabolism and transportation. The CD63 only interactome was enriched in Rab GTPases, SNARE proteins and sorting nexins while adding LMP1 into the interactome increased presence of signaling and ribosomal proteins. Our results showed that LMP1 alters the CD63 interactome, shifting the network of proteins enrichment from protein localization and vesicle mediated transportation to metabolic processes and translation. We also show that LMP1 interacts with mTor, Nedd4L and PP2A indicating formation of a multiprotein complex with CD63 thereby potentially regulating LMP1 dependent mTor signaling. Collectively, the comprehensive analysis of CD63 proximal interacting proteins provides insights into network of partners required for endocytic trafficking, extracellular vesicle cargo sorting, formation and secretion.

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