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Cis-Golgi cisternal assembly and biosynthetic activation occur sequentially in plants and algae.

ABSTRACT: The cisternal progression/maturation model of Golgi trafficking predicts that cis-Golgi cisternae are formed de novo on the cis-side of the Golgi. Here we describe structural and functional intermediates of the cis cisterna assembly process in high-pressure frozen algae (Scherffelia dubia, Chlamydomonas reinhardtii) and plants (Arabidopsis thaliana, Dionaea muscipula; Venus flytrap) as determined by electron microscopy, electron tomography and immuno-electron microscopy techniques. Our findings are as follows: (i) The cis-most (C1) Golgi cisternae are generated de novo from cisterna initiators produced by the fusion of 3-5 COPII vesicles in contact with a C2 cis cisterna. (ii) COPII vesicles fuel the growth of the initiators, which then merge into a coherent C1 cisterna. (iii) When a C1 cisterna nucleates its first cisterna initiator it becomes a C2 cisterna. (iv) C2-Cn cis cisternae grow through COPII vesicle fusion. (v) ER-resident proteins are recycled from cis cisternae to the ER via COPIa-type vesicles. (vi) In S. dubia the C2 cisternae are capable of mediating the self-assembly of scale protein complexes. (vii) In plants, ?90% of native ?-mannosidase I localizes to medial Golgi cisternae. (viii) Biochemical activation of cis cisternae appears to coincide with their conversion to medial cisternae via recycling of medial cisterna enzymes. We propose how the different cis cisterna assembly intermediates of plants and algae may actually be related to those present in the ERGIC and in the pre-cis Golgi cisterna layer in mammalian cells.

SUBMITTER: Donohoe BS

PROVIDER: S-EPMC3622843 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature

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Cis-Golgi cisternal assembly and biosynthetic activation occur sequentially in plants and algae.

Donohoe Bryon S BS Kang Byung-Ho BH Gerl Mathias J MJ Gergely Zachary R ZR McMichael Colleen M CM Bednarek Sebastian Y SY Staehelin L Andrew LA

Traffic (Copenhagen, Denmark) 20130225 5

The cisternal progression/maturation model of Golgi trafficking predicts that cis-Golgi cisternae are formed de novo on the cis-side of the Golgi. Here we describe structural and functional intermediates of the cis cisterna assembly process in high-pressure frozen algae (Scherffelia dubia, Chlamydomonas reinhardtii) and plants (Arabidopsis thaliana, Dionaea muscipula; Venus flytrap) as determined by electron microscopy, electron tomography and immuno-electron microscopy techniques. Our findings ...[more]

PMID: 23369235

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Project description:Rab6, the most abundant Golgi associated small GTPase, consists of 2 equally common isoforms, Rab6A and Rab6A', that differ in 3 amino acids and localize to trans Golgi cisternae. The two isoforms are largely redundant in function and hence are often referred to generically as Rab6. Rab6 loss-of-function inhibits retrograde Golgi trafficking, induces an increase in Golgi cisternal number in HeLa cells and delays the cell surface appearance of the anterograde cargo protein, VSVG. We hypothesized that these effects are linked and might be explained by a cisternal-specific delay in cargo transport. In pulse chase experiments using a deconvolved, confocal line scanning approach to score the distribution of the tsO45 mutant of VSVG protein in Rab6 depleted cells, we found that anterograde transport at 32 °C, permissive conditions, through the Golgi apparatus was locally delayed, almost tenfold, between medial and trans Golgi cisterna. Cis to medial transport was nearly normal as was trans Golgi to TGN transport. TGN exit was unaffected by Rab6 depletion. These effects were the same with either of two siRNAs. Similar intra-Golgi transport delays were seen at 37 °C with RUSH VSVG or a RUSH GPI-anchored construct using a biotin pulse to release the marker proteins from the ER. Using 3D-SIM, a super resolution approach, we found that RUSH VSVG transport was delayed pre-trans Golgi. These visual approaches suggest a selective slowing of anterograde transport relative to 3 different marker proteins downstream of the trans Golgi. Using a biochemical approach, we found that the onset of VSVG endoglycosidase H resistance in Rab6 depleted cells was delayed. Depletion of neither Rab6A or Rab6A' isoforms alone had any effect on anterograde transport through the Golgi suggesting that Rab6A and Rab6A' act coordinately. Delayed cargo transport conditions correlate strongly with a proliferation of Golgi cisternae observed in earlier electron microscopy. Our results strongly indicate that Rab6 is selectively required for rapid anterograde transport from the medial to trans Golgi. We suggest that the observed correlation with localized cisternal proliferation fits best with a cisternal progression model of Golgi function.

Dataset Information

Cis-Golgi cisternal assembly and biosynthetic activation occur sequentially in plants and algae.

Publications

Cis-Golgi cisternal assembly and biosynthetic activation occur sequentially in plants and algae.

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