A genetic and molecular characterization of the garnet gene of Drosophila melanogaster

Genome ◽  
1999 ◽  
Vol 42 (6) ◽  
pp. 1183-1193 ◽  
Author(s):  
Vett K Lloyd ◽  
D A Sinclair ◽  
R Wennberg ◽  
T S Warner ◽  
B M Honda ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Pigment Granule ◽  
Initial Step ◽  
Genetic Dissection ◽  
Trans Golgi Network ◽  
Intracellular Vesicle ◽  
Intracellular Sorting ◽  
Golgi Network ◽  
Delta Subunit

The garnet gene was one of the first genes to be identified in Drosophila melanogaster. Mutations in the garnet gene affect both of the biochemically distinct types of pigments in the eye and disrupt pigmentation of other organs. As an initial step in the analysis of this gene, we have analyzed the pigmentation defects in several of the garnet alleles. We have also cloned the gene and examined its expression in various tissues and at different stages of development. The garnet gene is expressed throughout development and in all tissues examined. Structurally related sequences can be detected in a variety of other eukaryotes. The predicted protein sequence of the garnet product resembles clathrin and nonclathrin adaptin proteins and is highly similar to the delta subunit of the newly isolated mammalian AP-3 adaptin complex, which is associated with the trans-Golgi network and endosomes. This suggests that garnet encodes a protein that acts in the intracellular sorting and trafficking of vesicles from the trans-Golgi network to endosomes, and related specialized organelles such as the pigment granule. This finding provides an explanation for the phenotype of garnet mutations and predicts that other Drosophila eye-colour genes will be a rich resource for the genetic dissection of intracellular vesicle transport.Key words: garnet, Drosophila melanogaster, AP-3, eye pigments.

scholarly journals Proteomic Characterization of Isolated Arabidopsis Clathrin-Coated Vesicles Reveals Evolutionarily Conserved and Plant Specific Components

2021 ◽  
Author(s):  
Dana A. Dahhan ◽  
Gregory D. Reynolds ◽  
Jessica J. Cárdenas ◽  
Dominique Eeckhout ◽  
Alexander Johnson ◽  
...  
Keyword(s):  
Secretory Pathway ◽  
Adaptor Protein ◽  
Coated Vesicles ◽  
Mass Spectrometry Analysis ◽  
Chemical Labeling ◽  
Spectrometry Analysis ◽  
Trans Golgi Network ◽  
Evolutionarily Conserved ◽  
Golgi Network

In eukaryotes, clathrin-coated vesicles (CCVs) facilitate the internalization of material from the cell surface as well as the movement of cargo in post-Golgi trafficking pathways. This diversity of functions is partially provided by multiple monomeric and multimeric clathrin adaptor complexes that provide compartment and cargo selectivity. The adaptor-protein AP-1 complex operates as part of the secretory pathway at the trans-Golgi network, while the AP-2 complex and the TPLATE complex (TPC) jointly operate at the plasma membrane to execute clathrin-mediated endocytosis. Key to our further understanding of clathrin-mediated trafficking in plants will be the comprehensive identification and characterization of the network of evolutionarily conserved and plant-specific core and accessory machinery involved in the formation and targeting of CCVs. To facilitate these studies, we have analyzed the proteome of enriched trans-Golgi network/early endosome-derived and endocytic CCVs isolated from dividing and expanding suspension-cultured Arabidopsis cells. Tandem mass spectrometry analysis results were validated by differential chemical labeling experiments to identify proteins co-enriching with CCVs. Proteins enriched in CCVs included previously characterized CCV components and cargos such as the vacuolar sorting receptors in addition to conserved and plant-specific components whose function in clathrin-mediated trafficking has not been previously defined. Notably, in addition to AP-1 and AP-2, all subunits of the AP-4 complex, but not AP-3 or AP-5, were found to be in high abundance in the CCV proteome. The association of AP-4 with suspension-cultured Arabidopsis CCVs is further supported via additional biochemical data.

scholarly journals Autoantigen Golgin-97, an Effector of Arl1 GTPase, Participates in Traffic from the Endosome to the Trans-Golgi Network

2004 ◽  
Vol 15 (10) ◽  
pp. 4426-4443 ◽  
Author(s):  
Lei Lu ◽  
Guihua Tai ◽  
Wanjin Hong
Keyword(s):  
Functional Study ◽  
Toxin B ◽  
Membrane Recruitment ◽  
Trans Golgi Network ◽  
Transport Assay ◽  
Retrograde Traffic ◽  
Golgi Network

The precise cellular function of Arl1 and its effectors, the GRIP domain Golgins, is not resolved, despite our recent understanding that Arl1 regulates the membrane recruitment of these Golgins. In this report, we describe our functional study of Golgin-97. Using a Shiga toxin B fragment (STxB)-based in vitro transport assay, we demonstrated that Golgin-97 plays a role in transport from the endosome to the trans-Golgi network (TGN). The recombinant GRIP domain of Golgin-97 as well as antibodies against Golgin-97 inhibited the transport of STxB in vitro. Membrane-associated Golgin-97, but not its cytosolic pool, was required in the in vitro transport assay. The kinetic characterization of inhibition by anti-Golgin-97 antibody in comparison with anti-Syntaxin 16 antibody established that Golgin-97 acts before Syntaxin 16 in endosome-to-TGN transport. Knock down of Golgin-97 or Arl1 by their respective small interference RNAs (siRNAs) also significantly inhibited the transport of STxB to the Golgi in vivo. In siRNA-treated cells with reduced levels of Arl1, internalized STxB was instead distributed peripherally. Microinjection of Golgin-97 antibody led to the fragmentation of Golgi apparatus and the arrested transport to the Golgi of internalized Cholera toxin B fragment. We suggest that Golgin-97 may function as a tethering molecule in endosome-to-TGN retrograde traffic.

Phosphoinositides and membrane traffic at the trans-Golgi network.

2005 ◽  
Vol 72 ◽  
pp. 31-38 ◽  
Author(s):  
Rawshan R. Choudhury ◽  
Noora Hyvola ◽  
Martin Lowe
Keyword(s):  
Plasma Membrane ◽  
Secretory Pathway ◽  
Binding Proteins ◽  
Molecular Mechanisms ◽  
Membrane Traffic ◽  
Trans Golgi Network ◽  
Cargo Proteins ◽  
Golgi Network

Cargo proteins moving along the secretory pathway are sorted at the TGN (trans-Golgi network) into distinct carriers for delivery to the plasma membrane or endosomes. Recent studies in yeast and mammals have shown that formation of these carriers is regulated by PtdIns(4)P. This phosphoinositide is abundant at the TGN and acts to recruit components required for carrier formation to the membrane. Other phosphoinositides are also present on the TGN, but the extent to which they regulate trafficking is less clear. Further characterization of phosphoinositide kinases and phosphatases together with identification of new TGN-associated phosphoinositide-binding proteins will reveal the extent to which different phosphoinositides regulate TGN trafficking, and help define the molecular mechanisms involved.

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