Endocytosis of cationized ferritin by coated vesicles of soybean protoplasts

107

We report on the sub-cellular localisation and function of m-Rabmc, a N-myristoylated plant-specific Rab-GTPase previously characterised at the molecular level and also by structural analysis in Mesembryanthemum crystallinum. By confocal laser scanning microscopy, we identified m-Rabmc predominantly on the prevacuolar compartment of the lytic vacuole but also on the Golgi apparatus in various plant cell types. Two complementary approaches were used immunocytochemistry and cyan fluorescent protein (CFP)/yellow fluorescent protein (YFP)-fusion proteins. Co-localisation studies of m-Rabmc with a salinity stress modulated integral calcium-ATPase suggest involvement of m-Rabmc in a plant-specific transport pathway to the prevacuolar compartment of the lytic vacuole. This hypothesis was strengthened by the inhibition of the transport of aleurain fused to green fluorescent protein (GFP), a marker of the lytic vacuole, in the presence of the dominant negative mutant m-Rabmc(N147I) in Arabidopsis thaliana protoplasts. The inhibitory effect of m-Rabmc(N147I) was specific for the transport pathway to the lytic vacuole, since the transport of chitinase-YFP, a marker for the neutral vacuole, was not hindered by the mutant.

Section: The Vacuolar Pathway Versus the Endocytic Pathwaymentioning

confidence: 93%

The N-myristoylated Rab-GTPase m-Rabmc is involved in post-Golgi trafficking events to the lytic vacuole in plant cells

Bolte

Brown

Satiat‐Jeunemaître

2004

107

“…We now also provide evidence, that a significant proportion of the PIN proteins is associated with the Golgi apparatus. This association suggests that, a level of regulation for PIN trafficking takes place at the Golgi, operating either as an exocytic and/or endocytic compartment (Tanchak et al, 1984;Bolte et al, 2004b;Hawes and Satiat-Jeunemaitre, 2005). These active PIN trafficking pathways between the Golgi stacks and plasma membrane do neither overlap with the vacuolar secretory pathway nor with the putative 'late' endocytic pathways (Tse et al, 2004) because no PVCs were PIN-labelled.…”

Section: Pin Protein Trafficking Involves the Golgi To Plasma Membranmentioning

confidence: 99%

The plasma membrane recycling pathway and cell polarity in plants: studies on PIN proteins

Boutté

Crosnier

Carraro

et al. 2006

137

136

Results AtPIN labelled PIN proteins in root cellsWestern blot analysis The purified AtPIN antiserum labelled one major band of approximately 68-70 kDa (Fig. 1), on western blots of wholeThe PIN-FORMED (PIN) proteins are plasma-membraneassociated facilitators of auxin transport. They are often targeted to one side of the cell only through subcellular mechanisms that remain largely unknown. Here, we have studied the potential roles of the cytoskeleton and endomembrane system in the localisation of PIN proteins. Immunocytochemistry and image analysis on root cells from Arabidopsis thaliana and maize showed that 10-30% of the intracellular PIN proteins mapped to the Golgi network, but never to prevacuolar compartments. The remaining 70-90% were associated with yet to be identified structures. The maintenance of PIN proteins at the plasma membrane depends on a BFA-sensitive machinery, but not on microtubules and actin filaments.The polar localisation of PIN proteins at the plasmamembrane was not reflected by any asymmetric distribution of cytoplasmic organelles. In addition, PIN proteins were inserted in a symmetrical manner at both sides of the cell plate during cytokinesis. Together, the data indicate that the localisation of PIN proteins is a postmitotic event, which depends on local characteristics of the plasma membrane and its direct environment. In this context, we present evidence that microtubule arrays might define essential positional information for PIN localisation. This information seems to require the presence of an intact cell wall.

“…In plants, only clathrin-dependent endocytosis has been described at the ultrastructural level in protoplasts using cationized ferritin and heavy metals (Tanchak et al, 1984;Fowke et al, 1991;Low and Chandra, 1994). Time-course analysis showed a degradation pathway by which the markers were first internalized in clathrin-coated vesicles at the PM and then delivered to partially coated reticulum, multivesicular bodies (MVBs) and finally to vacuoles.…”

Section: Introductionmentioning

confidence: 99%

Distinct endocytic pathways identified in tobacco pollen tubes using charged nanogold

Moscatelli

Ciampolini

Rodighiero

et al. 2007

129

198

In an attempt to dissect endocytosis in Nicotiana tabacum L. pollen tubes, two different probes – positively or negatively charged nanogold – were employed. The destiny of internalized plasma membrane domains, carrying negatively or positively charged residues, was followed at the ultrastructural level and revealed distinct endocytic pathways. Time-course experiments and electron microscopy showed internalization of subapical plasma-membrane domains that were mainly recycled to the secretory pathway through the Golgi apparatus and a second mainly degradative pathway involving plasma membrane retrieval at the tip. In vivo time-lapse experiments using FM4-64 combined with quantitative analysis confirmed the existence of distinct internalization regions. Ikarugamycin, an inhibitor of clathrin-dependent endocytosis, allowed us to further dissect the endocytic process: electron microscopy and time-lapse studies suggested that clathrin-dependent endocytosis occurs in the tip and subapical regions, because recycling of positively charged nanogold to the Golgi bodies and the consignment of negatively charged nanogold to vacuoles were affected. However, intact positively charged-nanogold transport to vacuoles supports the idea that an endocytic pathway that does not require clathrin is also present in pollen tubes.