The factors and mechanisms involved in vacuolar transport in plants, and in particular those directing vesicles to their target endomembrane compartment, remain largely unknown. To identify components of the vacuolar trafficking machinery, we searched for Arabidopsis modified transport to the vacuole (mtv) mutants that abnormally secrete the synthetic vacuolar cargo VAC2. We report here on the identification of 17 mtv mutations, corresponding to mutant alleles of MTV2/VSR4, MTV3/PTEN2A MTV7/EREL1, MTV8/ARFC1, MTV9/PUF2, MTV10/VPS3, MTV11/VPS15, MTV12/GRV2, MTV14/GFS10, MTV15/BET11, MTV16/VPS51, MTV17/VPS54, and MTV18/VSR1 Eight of the MTV proteins localize at the interface between the trans-Golgi network (TGN) and the multivesicular bodies (MVBs), supporting that the trafficking step between these compartments is essential for segregating vacuolar proteins from those destined for secretion. Importantly, the GARP tethering complex subunits MTV16/VPS51 and MTV17/VPS54 were found at endoplasmic reticulum (ER)- and microtubule-associated compartments (EMACs). Moreover, MTV16/VPS51 interacts with the motor domain of kinesins, suggesting that, in addition to tethering vesicles, the GARP complex may regulate the motors that transport them. Our findings unveil a previously uncharacterized compartment of the plant vacuolar trafficking pathway and support a role for microtubules and kinesins in GARP-dependent transport of soluble vacuolar cargo in plants.

• Klíčová slova
• microtubules, trafficking, vacuoles,
• MeSH
• alely MeSH
• Arabidopsis genetika   metabolismus   MeSH
• cytoplazmatické vezikuly genetika   metabolismus   MeSH
• endoplazmatické retikulum genetika   metabolismus   MeSH
• Golgiho aparát genetika   metabolismus   MeSH
• kineziny genetika   metabolismus   MeSH
• mikrotubuly genetika   metabolismus   MeSH
• multivezikulární tělíska genetika   metabolismus   MeSH
• mutace MeSH
• proteiny huseníčku genetika   metabolismus   MeSH
• transport proteinů genetika   MeSH
• vakuoly genetika   metabolismus   MeSH
• vezikulární transportní proteiny genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kineziny MeSH
• proteiny huseníčku MeSH
• vezikulární transportní proteiny MeSH

BACKGROUND: The Golgi apparatus is a central meeting point for the endocytic and exocytic systems in eukaryotic cells, and the organelle's dysfunction results in human disease. Its characteristic morphology of multiple differentiated compartments organized into stacked flattened cisternae is one of the most recognizable features of modern eukaryotic cells, and yet how this is maintained is not well understood. The Golgi is also an ancient aspect of eukaryotes, but the extent and nature of its complexity in the ancestor of eukaryotes is unclear. Various proteins have roles in organizing the Golgi, chief among them being the golgins. RESULTS: We address Golgi evolution by analyzing genome sequences from organisms which have lost stacked cisternae as a feature of their Golgi and those that have not. Using genomics and immunomicroscopy, we first identify Golgi in the anaerobic amoeba Mastigamoeba balamuthi. We then searched 87 genomes spanning eukaryotic diversity for presence of the most prominent proteins implicated in Golgi structure, focusing on golgins. We show some candidates as animal specific and others as ancestral to eukaryotes. CONCLUSIONS: None of the proteins examined show a phyletic distribution that correlates with the morphology of stacked cisternae, suggesting the possibility of stacking as an emergent property. Strikingly, however, the combination of golgins conserved among diverse eukaryotes allows for the most detailed reconstruction of the organelle to date, showing a sophisticated Golgi with differentiated compartments and trafficking pathways in the common eukaryotic ancestor.

• Klíčová slova
• GRASP, Golgi apparatus, evolutionary cell biology, golgin, membrane trafficking, secretion,
• MeSH
• Amoeba MeSH
• biologická evoluce * MeSH
• eukaryotické buňky fyziologie   MeSH
• fylogeneze * MeSH
• Golgiho aparát genetika   metabolismus   MeSH
• kultivované buňky MeSH
• transport proteinů fyziologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

The export of membrane proteins along the secretory pathway is initiated at the endoplasmic reticulum after proteins are folded and packaged inside this organelle by their recruiting into the coat complex COPII vesicles. It is proposed that cargo receptors are required for the correct transport of proteins to its target membrane, however, little is known about ER export signals for cargo receptors. Erv14/Cornichon belong to a well conserved protein family in Eukaryotes, and have been proposed to function as cargo receptors for many transmembrane proteins. Amino acid sequence alignment showed the presence of a conserved acidic motif in the C-terminal in homologues from plants and yeast. Here, we demonstrate that mutation of the C-terminal acidic motif from ScErv14 or OsCNIH1, did not alter the localization of these cargo receptors, however it modified the proper targeting of the plasma membrane transporters Nha1p, Pdr12p and Qdr2p. Our results suggest that mistargeting of these plasma membrane proteins is a consequence of a weaker interaction between the cargo receptor and cargo proteins caused by the mutation of the C-terminal acidic motif.

• Klíčová slova
• Acidic motif, Cargo selection, Cornichon, ScErv14,
• MeSH
• ABC transportéry genetika   MeSH
• aminokyselinové motivy genetika   MeSH
• buněčná membrána genetika   metabolismus   MeSH
• COP-vezikuly genetika   metabolismus   MeSH
• endoplazmatické retikulum genetika   metabolismus   MeSH
• Golgiho aparát genetika   metabolismus   MeSH
• membránové proteiny genetika   metabolismus   MeSH
• membránové transportní proteiny genetika   MeSH
• Na(+)-H(+) antiport genetika   MeSH
• rýže (rod) genetika   MeSH
• Saccharomyces cerevisiae - proteiny genetika   MeSH
• Saccharomyces cerevisiae genetika   MeSH
• sbalování proteinů MeSH
• sekvence aminokyselin genetika   MeSH
• sekvenční seřazení MeSH
• transport proteinů genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ABC transportéry MeSH
• Erv14 protein, S cerevisiae MeSH Prohlížeč
• membránové proteiny MeSH
• membránové transportní proteiny MeSH
• Na(+)-H(+) antiport MeSH
• NHX1 protein, S cerevisiae MeSH Prohlížeč
• Pdr12 protein, S cerevisiae MeSH Prohlížeč
• Qdr2 protein, S cerevisiae MeSH Prohlížeč
• Saccharomyces cerevisiae - proteiny MeSH