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BMP-regulated exosomes from Drosophila male reproductive glands reprogram female behavior.

Corrigan L, Redhai S, Leiblich A, Fan SJ, Perera SM, Patel R, Gandy C, Wainwright SM, Morris JF, Hamdy F, Goberdhan DC, Wilson C - J. Cell Biol. (2014)

Bottom Line: Male reproductive glands secrete signals into seminal fluid to facilitate reproductive success.Exosome release was required to inhibit female remating behavior, suggesting that exosomes are downstream effectors of BMP signaling.These results demonstrate a new function for the MVB-exosome pathway in the reproductive tract that appears to be conserved across evolution.

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Affiliation: Department of Physiology, Anatomy and Genetics and Nuffield Department of Surgical Sciences, University of Oxford, Oxford OX1 3QX, England, UK.

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SCs secrete CD63-GFP–positive, exosome-like puncta. (A–D) Paired accessory glands (AG) expressing CD63-GFP in SCs connect to the ejaculatory duct (ED; A). Images of different magnifications show surface sections of SCs within the AG epithelium (B; from square in A, arrows mark two SCs), a transverse section through the lumen (C; bracketed region in B, arrows mark two SCs), and CD63-GFP–positive puncta in the lumen (D; square in C; an image at higher confocal gain setting is shown in Fig. S1 F). (E and F) High magnification images of surface (E) and transverse (F; asterisk marks the lumen containing GFP puncta) sections through the SC show that CD63-GFP is also found at the apical plasma membrane (arrowhead; F) and the limiting membrane of SC vacuoles, the majority of which have a dense ANCE-positive core (highlighted with arrows). One or two CD63-GFP–lined compartments per SC lack an ANCE-stained core but contain fluorescent puncta (E, asterisk). Fasciclin3 (Fas3) marks septate junctions at the cell surface, and DAPI marks nuclei (note that SCs and MCs are binucleate). (G and H) Vital staining of SCs with LysoTracker red reveals CD63-GFP–positive puncta (putative ILVs; arrows) in large acidic compartments (mMVBLs), distinguishing them from other compartment classes, such as the more abundant secretory vacuoles (SVs) and immature late endosomes (iLEs). All images show SCs from 3-d-old males incubated at 28.5°C after eclosion. (I) Schematic representation of compartments within SC in H; values below indicate the mean total numbers of each compartment in 6-d-old control SCs counted from multiple sections along the apical–basal axis; the numbers of SCs analyzed are given in brackets. Approximate outline of the SC is marked in E–H. Bars: (A and B) 200 nm; (C) 20 µm; (D–H) 5 µm.
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fig1: SCs secrete CD63-GFP–positive, exosome-like puncta. (A–D) Paired accessory glands (AG) expressing CD63-GFP in SCs connect to the ejaculatory duct (ED; A). Images of different magnifications show surface sections of SCs within the AG epithelium (B; from square in A, arrows mark two SCs), a transverse section through the lumen (C; bracketed region in B, arrows mark two SCs), and CD63-GFP–positive puncta in the lumen (D; square in C; an image at higher confocal gain setting is shown in Fig. S1 F). (E and F) High magnification images of surface (E) and transverse (F; asterisk marks the lumen containing GFP puncta) sections through the SC show that CD63-GFP is also found at the apical plasma membrane (arrowhead; F) and the limiting membrane of SC vacuoles, the majority of which have a dense ANCE-positive core (highlighted with arrows). One or two CD63-GFP–lined compartments per SC lack an ANCE-stained core but contain fluorescent puncta (E, asterisk). Fasciclin3 (Fas3) marks septate junctions at the cell surface, and DAPI marks nuclei (note that SCs and MCs are binucleate). (G and H) Vital staining of SCs with LysoTracker red reveals CD63-GFP–positive puncta (putative ILVs; arrows) in large acidic compartments (mMVBLs), distinguishing them from other compartment classes, such as the more abundant secretory vacuoles (SVs) and immature late endosomes (iLEs). All images show SCs from 3-d-old males incubated at 28.5°C after eclosion. (I) Schematic representation of compartments within SC in H; values below indicate the mean total numbers of each compartment in 6-d-old control SCs counted from multiple sections along the apical–basal axis; the numbers of SCs analyzed are given in brackets. Approximate outline of the SC is marked in E–H. Bars: (A and B) 200 nm; (C) 20 µm; (D–H) 5 µm.

Mentions: Each AG secretes into a large lumen, which opens at its proximal end into the ejaculatory duct (Fig. 1, A–C). We expressed a GFP-tagged tetraspanin, CD63, a mammalian transmembrane exosome marker used previously to mark fly exosomes (Panáková et al., 2005; Gross, et al., 2012), independently in MCs and SCs (Figs. 1, A–C; and S1, A and B) using the GAL4/upstream activating sequence (UAS) system (Brand and Perrimon, 1993). In SCs of 3-d-old males, CD63-GFP localized to the limiting membrane of all large (>2-µm diameter) vacuoles and, in some cells, to the apical plasma membrane (Fig. 1 F). Not all vacuoles were identical, however; the majority appeared to be compartments previously described as secretory vacuoles (SVs; Rylett et al., 2007) because they had a dense core containing the secreted protease angiotensin-converting enzyme I (ANCE; Fig. 1, E and F). CD63-GFP–positive puncta, presumably representing intraluminal vesicles (ILVs), were also observed inside many vacuoles, especially in live specimens. These puncta were typically most prominent in at least one acidic compartment, stained with the vital acid-sensitive dye LysoTracker red (Fig. 1, G and H).


BMP-regulated exosomes from Drosophila male reproductive glands reprogram female behavior.

Corrigan L, Redhai S, Leiblich A, Fan SJ, Perera SM, Patel R, Gandy C, Wainwright SM, Morris JF, Hamdy F, Goberdhan DC, Wilson C - J. Cell Biol. (2014)

SCs secrete CD63-GFP–positive, exosome-like puncta. (A–D) Paired accessory glands (AG) expressing CD63-GFP in SCs connect to the ejaculatory duct (ED; A). Images of different magnifications show surface sections of SCs within the AG epithelium (B; from square in A, arrows mark two SCs), a transverse section through the lumen (C; bracketed region in B, arrows mark two SCs), and CD63-GFP–positive puncta in the lumen (D; square in C; an image at higher confocal gain setting is shown in Fig. S1 F). (E and F) High magnification images of surface (E) and transverse (F; asterisk marks the lumen containing GFP puncta) sections through the SC show that CD63-GFP is also found at the apical plasma membrane (arrowhead; F) and the limiting membrane of SC vacuoles, the majority of which have a dense ANCE-positive core (highlighted with arrows). One or two CD63-GFP–lined compartments per SC lack an ANCE-stained core but contain fluorescent puncta (E, asterisk). Fasciclin3 (Fas3) marks septate junctions at the cell surface, and DAPI marks nuclei (note that SCs and MCs are binucleate). (G and H) Vital staining of SCs with LysoTracker red reveals CD63-GFP–positive puncta (putative ILVs; arrows) in large acidic compartments (mMVBLs), distinguishing them from other compartment classes, such as the more abundant secretory vacuoles (SVs) and immature late endosomes (iLEs). All images show SCs from 3-d-old males incubated at 28.5°C after eclosion. (I) Schematic representation of compartments within SC in H; values below indicate the mean total numbers of each compartment in 6-d-old control SCs counted from multiple sections along the apical–basal axis; the numbers of SCs analyzed are given in brackets. Approximate outline of the SC is marked in E–H. Bars: (A and B) 200 nm; (C) 20 µm; (D–H) 5 µm.
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Related In: Results  -  Collection

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fig1: SCs secrete CD63-GFP–positive, exosome-like puncta. (A–D) Paired accessory glands (AG) expressing CD63-GFP in SCs connect to the ejaculatory duct (ED; A). Images of different magnifications show surface sections of SCs within the AG epithelium (B; from square in A, arrows mark two SCs), a transverse section through the lumen (C; bracketed region in B, arrows mark two SCs), and CD63-GFP–positive puncta in the lumen (D; square in C; an image at higher confocal gain setting is shown in Fig. S1 F). (E and F) High magnification images of surface (E) and transverse (F; asterisk marks the lumen containing GFP puncta) sections through the SC show that CD63-GFP is also found at the apical plasma membrane (arrowhead; F) and the limiting membrane of SC vacuoles, the majority of which have a dense ANCE-positive core (highlighted with arrows). One or two CD63-GFP–lined compartments per SC lack an ANCE-stained core but contain fluorescent puncta (E, asterisk). Fasciclin3 (Fas3) marks septate junctions at the cell surface, and DAPI marks nuclei (note that SCs and MCs are binucleate). (G and H) Vital staining of SCs with LysoTracker red reveals CD63-GFP–positive puncta (putative ILVs; arrows) in large acidic compartments (mMVBLs), distinguishing them from other compartment classes, such as the more abundant secretory vacuoles (SVs) and immature late endosomes (iLEs). All images show SCs from 3-d-old males incubated at 28.5°C after eclosion. (I) Schematic representation of compartments within SC in H; values below indicate the mean total numbers of each compartment in 6-d-old control SCs counted from multiple sections along the apical–basal axis; the numbers of SCs analyzed are given in brackets. Approximate outline of the SC is marked in E–H. Bars: (A and B) 200 nm; (C) 20 µm; (D–H) 5 µm.
Mentions: Each AG secretes into a large lumen, which opens at its proximal end into the ejaculatory duct (Fig. 1, A–C). We expressed a GFP-tagged tetraspanin, CD63, a mammalian transmembrane exosome marker used previously to mark fly exosomes (Panáková et al., 2005; Gross, et al., 2012), independently in MCs and SCs (Figs. 1, A–C; and S1, A and B) using the GAL4/upstream activating sequence (UAS) system (Brand and Perrimon, 1993). In SCs of 3-d-old males, CD63-GFP localized to the limiting membrane of all large (>2-µm diameter) vacuoles and, in some cells, to the apical plasma membrane (Fig. 1 F). Not all vacuoles were identical, however; the majority appeared to be compartments previously described as secretory vacuoles (SVs; Rylett et al., 2007) because they had a dense core containing the secreted protease angiotensin-converting enzyme I (ANCE; Fig. 1, E and F). CD63-GFP–positive puncta, presumably representing intraluminal vesicles (ILVs), were also observed inside many vacuoles, especially in live specimens. These puncta were typically most prominent in at least one acidic compartment, stained with the vital acid-sensitive dye LysoTracker red (Fig. 1, G and H).

Bottom Line: Male reproductive glands secrete signals into seminal fluid to facilitate reproductive success.Exosome release was required to inhibit female remating behavior, suggesting that exosomes are downstream effectors of BMP signaling.These results demonstrate a new function for the MVB-exosome pathway in the reproductive tract that appears to be conserved across evolution.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology, Anatomy and Genetics and Nuffield Department of Surgical Sciences, University of Oxford, Oxford OX1 3QX, England, UK.

Show MeSH
Related in: MedlinePlus