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Stepping stone: a cytohesin adaptor for membrane cytoskeleton restraint in the syncytial Drosophila embryo.

Liu J, Lee DM, Yu CG, Angers S, Harris TJ - Mol. Biol. Cell (2014)

Bottom Line: Elevating Sstn furrow levels had no effect on the steppke phenotype, but elevating Steppke furrow levels reversed the sstn phenotype, suggesting that Steppke acts downstream of Sstn and that additional mechanisms can recruit Steppke to furrows.Finally, the coiled-coil domain of Steppke was required for Sstn binding and in addition homodimerization, and its removal disrupted Steppke furrow localization and activity in vivo.Overall we propose that Sstn acts as a cytohesin adaptor that promotes Steppke activity for localized membrane cytoskeleton restraint in the syncytial Drosophila embryo.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Systems Biology, University of Toronto, Toronto, ON M5S 3G5, Canada.

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Step overexpression can overcome the effects of Sstn loss but not vice versa. (A) step shRNA had no apparent effect on GFP-Sstn localization to furrow membranes (arrows), and the localized GFP-Sstn had no apparent effect on the membrane expansion that occurs with Step loss (compare Amph staining). (B) sstn shRNA had no apparent effect on GFP-Step enrichment at basal furrow tips (arrows). (C) GFP-Step expression eliminated the furrow expansion that occurs with Sstn loss (quantified in graph as in Figure 5). Sporadic furrow loss is also observed in GFP-Step embryos that progress to early cellularization (arrows; 15 of 27 embryos were imaged with missing or shorter furrows at this stage) and was similarly seen in embryos with both GFP-Step and sstn shRNA expression (arrows; 22 of 52 embryos with missing or shorter furrows were imaged).
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Figure 7: Step overexpression can overcome the effects of Sstn loss but not vice versa. (A) step shRNA had no apparent effect on GFP-Sstn localization to furrow membranes (arrows), and the localized GFP-Sstn had no apparent effect on the membrane expansion that occurs with Step loss (compare Amph staining). (B) sstn shRNA had no apparent effect on GFP-Step enrichment at basal furrow tips (arrows). (C) GFP-Step expression eliminated the furrow expansion that occurs with Sstn loss (quantified in graph as in Figure 5). Sporadic furrow loss is also observed in GFP-Step embryos that progress to early cellularization (arrows; 15 of 27 embryos were imaged with missing or shorter furrows at this stage) and was similarly seen in embryos with both GFP-Step and sstn shRNA expression (arrows; 22 of 52 embryos with missing or shorter furrows were imaged).

Mentions: To investigate the pathway relationship between Sstn and Step, we tested how they affect each other's localization and RNA interference (RNAi) phenotypes. For GFP-Sstn, we observed similar local levels of the protein at the base of furrows in control embryos and in step RNAi embryos with abnormal perpendicular expansion of furrow tips (Figure 7A). Thus it seems that Step is not essential for the localization of overexpressed Sstn. Moreover, the Sstn overexpression could not overcome the effects of Step loss. Sstn overexpression also had no noticeable effect on otherwise genetically wild-type syncytial embryos.


Stepping stone: a cytohesin adaptor for membrane cytoskeleton restraint in the syncytial Drosophila embryo.

Liu J, Lee DM, Yu CG, Angers S, Harris TJ - Mol. Biol. Cell (2014)

Step overexpression can overcome the effects of Sstn loss but not vice versa. (A) step shRNA had no apparent effect on GFP-Sstn localization to furrow membranes (arrows), and the localized GFP-Sstn had no apparent effect on the membrane expansion that occurs with Step loss (compare Amph staining). (B) sstn shRNA had no apparent effect on GFP-Step enrichment at basal furrow tips (arrows). (C) GFP-Step expression eliminated the furrow expansion that occurs with Sstn loss (quantified in graph as in Figure 5). Sporadic furrow loss is also observed in GFP-Step embryos that progress to early cellularization (arrows; 15 of 27 embryos were imaged with missing or shorter furrows at this stage) and was similarly seen in embryos with both GFP-Step and sstn shRNA expression (arrows; 22 of 52 embryos with missing or shorter furrows were imaged).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4325841&req=5

Figure 7: Step overexpression can overcome the effects of Sstn loss but not vice versa. (A) step shRNA had no apparent effect on GFP-Sstn localization to furrow membranes (arrows), and the localized GFP-Sstn had no apparent effect on the membrane expansion that occurs with Step loss (compare Amph staining). (B) sstn shRNA had no apparent effect on GFP-Step enrichment at basal furrow tips (arrows). (C) GFP-Step expression eliminated the furrow expansion that occurs with Sstn loss (quantified in graph as in Figure 5). Sporadic furrow loss is also observed in GFP-Step embryos that progress to early cellularization (arrows; 15 of 27 embryos were imaged with missing or shorter furrows at this stage) and was similarly seen in embryos with both GFP-Step and sstn shRNA expression (arrows; 22 of 52 embryos with missing or shorter furrows were imaged).
Mentions: To investigate the pathway relationship between Sstn and Step, we tested how they affect each other's localization and RNA interference (RNAi) phenotypes. For GFP-Sstn, we observed similar local levels of the protein at the base of furrows in control embryos and in step RNAi embryos with abnormal perpendicular expansion of furrow tips (Figure 7A). Thus it seems that Step is not essential for the localization of overexpressed Sstn. Moreover, the Sstn overexpression could not overcome the effects of Step loss. Sstn overexpression also had no noticeable effect on otherwise genetically wild-type syncytial embryos.

Bottom Line: Elevating Sstn furrow levels had no effect on the steppke phenotype, but elevating Steppke furrow levels reversed the sstn phenotype, suggesting that Steppke acts downstream of Sstn and that additional mechanisms can recruit Steppke to furrows.Finally, the coiled-coil domain of Steppke was required for Sstn binding and in addition homodimerization, and its removal disrupted Steppke furrow localization and activity in vivo.Overall we propose that Sstn acts as a cytohesin adaptor that promotes Steppke activity for localized membrane cytoskeleton restraint in the syncytial Drosophila embryo.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Systems Biology, University of Toronto, Toronto, ON M5S 3G5, Canada.

Show MeSH