FM‐dyes as experimental probes for dissecting vesicle trafficking in living plant cells

Bolte, Susanne; Talbot, Christopher J.; Boutté, Yohann; Catrice, Olivier; Read, Nick D.; Satiat‐Jeunemaître, Béatrice

doi:10.1111/j.0022-2720.2004.01348.x

Cited by 528 publications

(512 citation statements)

References 59 publications

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“…Tse et al (2004) proposed that the PVC lies on the secretory and endocytic pathways, based on the analysis of FM4-64 internalization in tobacco BY-2 cells. However, uptake and distribution of FM4-64 might not reflect the genuine trafficking of plasma membrane proteins in plant cells (Bolte et al, 2004b). Our data provide compelling evidence that endocytosed PM proteins as well as cargos of the secretory pathway merge at the PVC/MVB.…”

Section: Discussionmentioning

confidence: 79%

Evidence for a sorting endosome in Arabidopsis root cells

et al. 2007

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SummaryIn eukaryotic cells, the endocytic and secretory pathways are key players in several physiological processes. These pathways are largely inter-connected in animal and yeast cells through organelles named sorting endosomes. Sorting endosomes are multi-vesicular compartments that redirect proteins towards various destinations, such as the lysosomes or vacuoles for degradation, the trans-Golgi network for retrograde transport and the plasma membrane for recycling. In contrast, cross-talk between the endocytic and secretory pathways has not been clearly established in plants, especially in terms of cargo protein trafficking. Here we show by co-localization analyses that endosomes labelled with the AtSORTING NEXIN1 (AtSNX1) protein overlap with the pre-vacuolar compartment in Arabidopsis root cells. In addition, alteration of the routing functions of AtSNX1 endosomes by drug treatments leads to mis-routing of endocytic and secretory cargo proteins. Based on these results, we propose that the AtSNX1 endosomal compartment represents a sorting endosome in root cells, and that this specialized organelle is conserved throughout eukaryotes.

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Section: Discussionmentioning

confidence: 79%

Evidence for a sorting endosome in Arabidopsis root cells

et al. 2007

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“…In plant cells, FM4-64 becomes localized to the Golgi, PVC and vacuolar membrane after 30 min of staining, and to many organelles involved in the secretory and recycling pathways, as well as the endocytic pathway, by 60 min (ref. 43). In yeast, secretion of endocytosed FM4-64 via the recycling pathway is more rapid, and around half of the endocytosed FM4-64 is secreted within 10 min via the recycling pathway 44 , probably through the TGN compartments.…”

Section: Discussionmentioning

confidence: 99%

Bifurcation of the endocytic pathway into Rab5-dependent and -independent transport to the vacuole

et al. 2014

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The yeast Rab5 homologue, Vps21p, is known to be involved both in the vacuolar protein sorting (VPS) pathway from the trans-Golgi network to the vacuole, and in the endocytic pathway from the plasma membrane to the vacuole. However, the intracellular location at which these two pathways converge remains unclear. In addition, the endocytic pathway is not completely blocked in yeast cells lacking all Rab5 genes, suggesting the existence of an unidentified route that bypasses the Rab5-dependent endocytic pathway. Here we show that convergence of the endocytic and VPS pathways occurs upstream of the requirement for Vps21p in these pathways. We also identify a previously unidentified endocytic pathway mediated by the AP-3 complex. Importantly, the AP-3-mediated pathway appears mostly intact in Rab5-disrupted cells, and thus works as an alternative route to the vacuole/ lysosome. We propose that the endocytic traffic branches into two routes to reach the vacuole: a Rab5-dependent VPS pathway and a Rab5-independent AP-3-mediated pathway.

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“…To test this, FM4-64, a water-soluble and membraneselective fluorescent dye frequently used as a membrane marker to monitor endocytosis in mammalian, fungal and plant cells [61], was used to investigate the potential involvement of PIP5K2 in vesicle trafficking. As shown in Figure 5A, after staining for 30 min, the FM4-64 dye was internalized, and a substantial number of fluorescent vesicles were detected in the cytosol of WT root cells.…”

Section: Vesicle Trafficking Is Decelerated In Pip5k2 Which Can Be Rmentioning

confidence: 99%

Arabidopsis phosphatidylinositol monophosphate 5-kinase 2 is involved in root gravitropism through regulation of polar auxin transport by affecting the cycling of PIN proteins

et al. 2011

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Phosphatidylinositol monophosphate 5-kinase (PIP5K) catalyzes the synthesis of PI-4,5-bisphosphate (PtdIns(4,5) P 2 ) by phosphorylation of PI-4-phosphate at the 5 position of the inositol ring, and is involved in regulating multiple developmental processes and stress responses. We here report on the functional characterization of Arabidopsis PIP5K2, which is expressed during lateral root initiation and elongation, and whose expression is enhanced by exogenous auxin. The knockout mutant pip5k2 shows reduced lateral root formation, which could be recovered with exogenous auxin, and interestingly, delayed root gravity response that could not be recovered with exogenous auxin. Crossing with the DR5-GUS marker line and measurement of free IAA content confirmed the reduced auxin accumulation in pip5k2. In addition, analysis using the membrane-selective dye FM4-64 revealed the decelerated vesicle trafficking caused by PtdIns(4,5)P 2 reduction, which hence results in suppressed cycling of PIN proteins (PIN2 and 3), and delayed redistribution of PIN2 and auxin under gravistimulation in pip5k2 roots. On the contrary, PtdIns(4,5) P 2 significantly enhanced the vesicle trafficking and cycling of PIN proteins. These results demonstrate that PIP5K2 is involved in regulating lateral root formation and root gravity response, and reveal a critical role of PIP5K2/PtdIns(4,5)P 2 in root development through regulation of PIN proteins, providing direct evidence of crosstalk between the phosphatidylinositol signaling pathway and auxin response, and new insights into the control of polar auxin transport.

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FM‐dyes as experimental probes for dissecting vesicle trafficking in living plant cells

Cited by 528 publications

References 59 publications

Evidence for a sorting endosome in Arabidopsis root cells

Evidence for a sorting endosome in Arabidopsis root cells

Bifurcation of the endocytic pathway into Rab5-dependent and -independent transport to the vacuole

Arabidopsis phosphatidylinositol monophosphate 5-kinase 2 is involved in root gravitropism through regulation of polar auxin transport by affecting the cycling of PIN proteins

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