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Microtubules provide directional information for core PCP function.

Matis M, Russler-Germain DA, Hu Q, Tomlin CJ, Axelrod JD - Elife (2014)

Bottom Line: Consistent with previous results, we find that the Ft/Ds/Fj-module has an effect on a MT-cytoskeleton.We show Ft/Ds/Fj-dependent initial polarization of the apical MT-cytoskeleton prior to global alignment of the core-module, reveal that the anchoring of apical non-centrosomal MTs at apical junctions is polarized, observe that directional trafficking of vesicles containing Dsh depends on Ft, and demonstrate the feasibility of this model by mathematical simulation.Together, these results support the hypothesis that Ft/Ds/Fj provides a signal to orient core PCP function via MT polarization.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, United States matism@uni-muenster.de.

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Vesicle identification and tubulin staining.(A) 24 hAPF Dsh::GFP wing co-labeled for Sec5 and Rab5. Many Dsh::GFP vesicles do not label with Sec5 or Rab5 (pink arrowheads), while some stain for Sec5 (yellow arrowhead) or Rab5 (blue arrowhead). (B) Tubulin staining in fz and dsh1 mutant flies. Compare to Figure 1—figure supplement 1C.DOI:http://dx.doi.org/10.7554/eLife.02893.018
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fig4s1: Vesicle identification and tubulin staining.(A) 24 hAPF Dsh::GFP wing co-labeled for Sec5 and Rab5. Many Dsh::GFP vesicles do not label with Sec5 or Rab5 (pink arrowheads), while some stain for Sec5 (yellow arrowhead) or Rab5 (blue arrowhead). (B) Tubulin staining in fz and dsh1 mutant flies. Compare to Figure 1—figure supplement 1C.DOI:http://dx.doi.org/10.7554/eLife.02893.018

Mentions: Distally biased microtubule (MT)-dependent trafficking of Fz-containing vesicles has been shown to occur during polarization of the core PCP proteins, and both are sensitive to MT disruption, suggesting that transport is required for polarization (Shimada et al., 2006). Since we have proposed that Dsh is the critical determinant that must be asymmetrically localized (Amonlirdviman et al., 2005; Axelrod, 2009), we also examined Dsh::GFP vesicle movement in developing wings in the AJ plane, between 15 and 32 hr after puparium formation (hAPF). The majority of vesicles (80%, n = 1192) moved along the P/D axis, and showed a significant though modest bias towards distal vs proximal transport (Figure 4A,C–D; Video 2). Dsh::GFP vesicles exhibited two distinct patterns of trafficking. First, and most commonly, vesicles emerged from one side of the cell and were transported directly across the length of the cell to be incorporated into the membrane of an opposing cellular face (Figure 4A′; Video 2). Movement was highly linear and processive, though occasional backtracking and zig-zagging was observed. Often, multiple vesicles followed similar paths in a given cell, with vesicle scission and fusion appearing to occur repeatedly at specific sites. Second, a minority of Dsh::GFP vesicles took staggered paths without directional bias, paused frequently, and often left the apical plane of the cell (Figure 4B,B′; Video 3). The former pattern likely reflects polarized transcytosis, resulting in net transport of Dsh to the distal membrane, while the latter reflects a recycling pathway. Consistently, in fixed specimens, only a minor fraction of Dsh vesicles co-stains with the early endosome marker Rab5 or exocyst protein Sec5 (Figure 4—figure supplement 1A). Thus, a minority of vesicles moves through the recycling pathway while the majority of Dsh vesicles appears to be part of a transcytosis pathway. In contrast, we see no directionally biased trafficking of Vang::YFP vesicles (Figure 4E). Note that biased directional transport of any one component of the core PCP proteins should be sufficient to provide an input bias; bulk transport to achieve the remainder of asymmetric localization is expected to occur by diffusion in combination with feedback at intercellular junctions. Together with prior data, our observations suggest that the ‘distal’ components Fz and Dsh, but not ‘proximal’ components, are subject to directional trafficking.10.7554/eLife.02893.017Figure 4.Dsh::GFP vesicles move preferentially toward the distal side of the cell.


Microtubules provide directional information for core PCP function.

Matis M, Russler-Germain DA, Hu Q, Tomlin CJ, Axelrod JD - Elife (2014)

Vesicle identification and tubulin staining.(A) 24 hAPF Dsh::GFP wing co-labeled for Sec5 and Rab5. Many Dsh::GFP vesicles do not label with Sec5 or Rab5 (pink arrowheads), while some stain for Sec5 (yellow arrowhead) or Rab5 (blue arrowhead). (B) Tubulin staining in fz and dsh1 mutant flies. Compare to Figure 1—figure supplement 1C.DOI:http://dx.doi.org/10.7554/eLife.02893.018
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4151085&req=5

fig4s1: Vesicle identification and tubulin staining.(A) 24 hAPF Dsh::GFP wing co-labeled for Sec5 and Rab5. Many Dsh::GFP vesicles do not label with Sec5 or Rab5 (pink arrowheads), while some stain for Sec5 (yellow arrowhead) or Rab5 (blue arrowhead). (B) Tubulin staining in fz and dsh1 mutant flies. Compare to Figure 1—figure supplement 1C.DOI:http://dx.doi.org/10.7554/eLife.02893.018
Mentions: Distally biased microtubule (MT)-dependent trafficking of Fz-containing vesicles has been shown to occur during polarization of the core PCP proteins, and both are sensitive to MT disruption, suggesting that transport is required for polarization (Shimada et al., 2006). Since we have proposed that Dsh is the critical determinant that must be asymmetrically localized (Amonlirdviman et al., 2005; Axelrod, 2009), we also examined Dsh::GFP vesicle movement in developing wings in the AJ plane, between 15 and 32 hr after puparium formation (hAPF). The majority of vesicles (80%, n = 1192) moved along the P/D axis, and showed a significant though modest bias towards distal vs proximal transport (Figure 4A,C–D; Video 2). Dsh::GFP vesicles exhibited two distinct patterns of trafficking. First, and most commonly, vesicles emerged from one side of the cell and were transported directly across the length of the cell to be incorporated into the membrane of an opposing cellular face (Figure 4A′; Video 2). Movement was highly linear and processive, though occasional backtracking and zig-zagging was observed. Often, multiple vesicles followed similar paths in a given cell, with vesicle scission and fusion appearing to occur repeatedly at specific sites. Second, a minority of Dsh::GFP vesicles took staggered paths without directional bias, paused frequently, and often left the apical plane of the cell (Figure 4B,B′; Video 3). The former pattern likely reflects polarized transcytosis, resulting in net transport of Dsh to the distal membrane, while the latter reflects a recycling pathway. Consistently, in fixed specimens, only a minor fraction of Dsh vesicles co-stains with the early endosome marker Rab5 or exocyst protein Sec5 (Figure 4—figure supplement 1A). Thus, a minority of vesicles moves through the recycling pathway while the majority of Dsh vesicles appears to be part of a transcytosis pathway. In contrast, we see no directionally biased trafficking of Vang::YFP vesicles (Figure 4E). Note that biased directional transport of any one component of the core PCP proteins should be sufficient to provide an input bias; bulk transport to achieve the remainder of asymmetric localization is expected to occur by diffusion in combination with feedback at intercellular junctions. Together with prior data, our observations suggest that the ‘distal’ components Fz and Dsh, but not ‘proximal’ components, are subject to directional trafficking.10.7554/eLife.02893.017Figure 4.Dsh::GFP vesicles move preferentially toward the distal side of the cell.

Bottom Line: Consistent with previous results, we find that the Ft/Ds/Fj-module has an effect on a MT-cytoskeleton.We show Ft/Ds/Fj-dependent initial polarization of the apical MT-cytoskeleton prior to global alignment of the core-module, reveal that the anchoring of apical non-centrosomal MTs at apical junctions is polarized, observe that directional trafficking of vesicles containing Dsh depends on Ft, and demonstrate the feasibility of this model by mathematical simulation.Together, these results support the hypothesis that Ft/Ds/Fj provides a signal to orient core PCP function via MT polarization.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, United States matism@uni-muenster.de.

Show MeSH
Related in: MedlinePlus