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. 2014 Mar 3;204(5):793–806. doi: 10.1083/jcb.201309084

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© 2014 Chi et al.

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Figure 7. — Loss of Vps1 dynamin family GTPase results in deficient fission of SNX-BAR–coated tubules. (A) Subcellular fractionation analysis of Vps10-myc. Spheroplasts derived from the indicated strains were lysed and a low-speed membrane pellet (13,000 g) and a high-speed membrane pellet (100,000 g) were generated. Equivalent amounts of each fraction were separated by SDS-PAGE and immunoblotted with anti-myc antibody. The position of full-length Vps10-myc is indicated, as well as degradative products that arise as a result of proteolysis by vacuolar proteases. The positions of molecular mass (kD) markers are indicated. (B) Time-lapse gallery of Mvp1-GFP– and Vps17-coated endosomes in wild-type and vps1Δ cells by time-lapse fluorescence deconvolution microscopy. Arrows highlight a tubule of interest in each time-lapse series and open arrow (<) indicates a fission event. (C) Tubule lifetimes were calculated as described in the text for Mvp1-GFP– or Vps17-GFP–coated tubules. Vps17 tubule lifetimes were significantly increased in vps1Δ cells (P = 0.0005) and in mvp1Δ cells (P = 0.043). (D) Vps17 tubule frequencies were calculated as described in the text. The frequency of Vps17-GFP tubules increased approximately twofold in mvp1Δ cells (P < 0.0001).