1. Volume 4, Issue 5, Pages 845-853 (September 2011)
Vacuolar Sorting Receptor (VSR) Proteins Reach the Plasma Membrane in Germinating Pollen Tubes 
Wang Hao , Zhuang Xiao-Hong , Hillmer Stefan , Robinson David G. , Jiang Li-Wen  
Molecular Plant 
Volume 4, Issue 5, Pages (September 2011)
DOI: /mp/ssr011
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

2. Figure 1
Immunofluorescent Detection of VSR Proteins in Germinating Lily and Tobacco Pollen Tubes.
(A, B) Germinated lily pollen tubes were chemically fixed and immunolabeled with VSRat-1 and BP-80 CT antibodies, respectively, followed by confocal imaging.
(C, D) Germinated lily pollen tubes were first treated with wortmannin (Wort) at 16.5 μM for 1 h, followed by chemical fixation before labeling with VSRat-1 and BP-80 CT antibodies, respectively, followed by confocal imaging. Arrows indicate examples of wortmannin-induced enlarged PVCs.
(E, F) Germinated tobacco pollen tubes were chemically fixed and immunolabeled with VSRat-1 and BP-80 CT antibodies, respectively, followed by confocal imaging.
(G, H) Germinated tobacco pollen tubes were first treated with wortmannin (Wort) at 16.5 μM for 1 h, followed by chemical fixation before labeling with VSRat-1 and BP-80 CT antibodies, respectively, followed by confocal imaging. Arrows indicate examples of wortmannin-induced enlarged PVCs. DIC, differential interference contrast. Scale bar = 50 μm in (A–D) and 12.5 μm in (E–H).
Molecular Plant 2011 4, DOI: ( /mp/ssr011)
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3. Figure 2
Immunofluorescent Detection of SCAMP Proteins in Germinating Lily and Tobacco Pollen Tubes.
Untreated (A, B) or BFA-treated (at 10 μg ml−1 for 1 h) (C) germinated lily pollen tubes were chemically fixed and immunolabeled with SCAMP1 antibodies. (A) and (C) show confocal images collected from a single optical section, while (B) shows a 3-D projection of multiple sections. Untreated (D, E) or BFA-treated (at 10 μg ml−1 for 1 h) (F) germinated tobacco pollen tubes were chemically fixed and immunolabeled with SCAMP1 antibodies. (A) and (C) show confocal images collected from a single optical section, while (B) shows a 3-D projection of multiple sections. DIC, differential interference contrast. Scale bar = 25 μm in (A–C) and 12.5 μm in (D–F).
Molecular Plant 2011 4, DOI: ( /mp/ssr011)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

4. Figure 3
Double-Immunofluorescent Localization of VSR and SCAMP1 in Germinating Lily Pollen Tubes.
Germinated lily pollen tubes were fixed and labeled with antibodies of VSRat-1, BP-80 CT, and SCAMP1 (A, B) or SCAMP1 and Man1 (C), followed by confocal imaging. Germinated tobacco pollen tubes were fixed and labeled with antibodies of VSRat-1, BP-80 CT, and SCAMP1 (D, E) or SCAMP1 and Man1 (F), followed by confocal imaging. DIC, differential interference contrast. Scale bar = 50 μm in (A–D) or 12.5 μm in (E, F).
Molecular Plant 2011 4, DOI: ( /mp/ssr011)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions

5. Figure 4
Immunogold EM Localization of VSR and SCAMP on PM in Lily Pollen Tubes.
Ultrathin sections prepared from high-pressure freezing/frozen-substitute pollen samples were labeled with VSRat-1, BP-80 CT, and SCAMP1 antibodies. An overview of pollen tube was shown in (A) and arrows indicate examples of gold particles on the plasma membrane with antibodies of VSRat-1 (B), BP-80 CT (C), and SCAMP1 (D). Scale bar = 5 μm in (A), while, in (B–D), it is 500 nm.
Molecular Plant 2011 4, DOI: ( /mp/ssr011)
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6. Figure 5
Fusion of VSR-Positive Organelle with PM in Growing Tobacco Pollen Tube.
Germinating tobacco pollen tube co-expressing RFP–AtVSR2 and GFP–AtSCAMP4, under the control of the pollen-specific promoter Lat 52 (A), was subjected to 3-D confocal imaging using a Spinning Disc Confocal Microscopy (B). (C) Time-lapse collections of 3-D images of region (indicated by dashed box) in (B), arrowheads indicate examples of possible VSR–PM fusion in growing pollen tube, where the inserted boxes are the enlarged images of the indicated area. Scale bar in (B, C) is 25 nm.
Molecular Plant 2011 4, DOI: ( /mp/ssr011)
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7. Figure 6 VSR Dynamics and Possible Functions in Growing Pollen Tube.
Shown is a working model of possible sub-cellular localization, dynamics, and functions of VSR proteins in germinating pollen tube. SCAMP is highly enriched in the PM and apical region of the pollen tube where VSR is missing. In addition to a possible ER–Golgi–TGN–PVC/MVB–vacuole transport pathway, VSR/BP-80 could also reach the plasma membrane (PM) likely from the trans-Golgi network (TGN) or likely from the multivesicular body (MVB) because VSR was also found in PM in this study. VSR in PM could also internalize to reach TGN/EE for another round of sorting for PM delivery of soluble cargo proteins.
Molecular Plant 2011 4, DOI: ( /mp/ssr011)
Copyright © 2011 The Authors. All rights reserved. Terms and Conditions