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The Arf GAP SMAP2 is necessary for organized vesicle budding from the trans-Golgi network and subsequent acrosome formation in spermiogenesis.

Funaki T, Kon S, Tanabe K, Natsume W, Sato S, Shimizu T, Yoshida N, Wong WF, Ogura A, Ogawa T, Inoue K, Ogonuki N, Miki H, Mochida K, Endoh K, Yomogida K, Fukumoto M, Horai R, Iwakura Y, Ito C, Toshimori K, Watanabe T, Satake M - Mol. Biol. Cell (2013)

Bottom Line: In the present study, SMAP2 is detected on the TGN in the pachytene spermatocyte to the round spermatid stages of spermatogenesis.Furthermore, syntaxin2, a component of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex, is not properly concentrated at the site of acrosome formation.Thus this study reveals a link between SMAP2 and CALM/syntaxin2 in clathrin-coated vesicle formation from the TGN and subsequent acrosome formation.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Immunology, Department of Pathology, Institute of Development, Aging and Cancer, Tohoku University, Sendai 980-8575, Japan.

ABSTRACT
The trans-Golgi network (TGN) functions as a hub organelle in the exocytosis of clathrin-coated membrane vesicles, and SMAP2 is an Arf GTPase-activating protein that binds to both clathrin and the clathrin assembly protein (CALM). In the present study, SMAP2 is detected on the TGN in the pachytene spermatocyte to the round spermatid stages of spermatogenesis. Gene targeting reveals that SMAP2-deficient male mice are healthy and survive to adulthood but are infertile and exhibit globozoospermia. In SMAP2-deficient spermatids, the diameter of proacrosomal vesicles budding from TGN increases, TGN structures are distorted, acrosome formation is severely impaired, and reorganization of the nucleus does not proceed properly. CALM functions to regulate vesicle sizes, and this study shows that CALM is not recruited to the TGN in the absence of SMAP2. Furthermore, syntaxin2, a component of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex, is not properly concentrated at the site of acrosome formation. Thus this study reveals a link between SMAP2 and CALM/syntaxin2 in clathrin-coated vesicle formation from the TGN and subsequent acrosome formation. SMAP2-deficient mice provide a model for globozoospermia in humans.

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Electron microscopic observations of proacrosomal vesicle formation at the Golgi phase. (A) Germ cells from SMAP2(+/+) and (−/−) mice were processed for electron microscopic observations. C, cis-Golgi network; G, Golgi apparatus; T, trans-Golgi network. Bars, 1 μm. (B) Size distribution of proacrosomal vesicles observed in SMAP2(+/+) (closed bars) and (−/−) (open bars) germ cells at the Golgi phase. Using the images shown in A, we measured diameters for each proacrosomal vesicle. The areas of the vesicles were calculated using image-processing software and divided by the square of the magnification. The diameters are presented in relative units. Vesicle sizes are binned as indicated. See Supplemental Figure S3 for the detailed size distribution of vesicles.
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Figure 5: Electron microscopic observations of proacrosomal vesicle formation at the Golgi phase. (A) Germ cells from SMAP2(+/+) and (−/−) mice were processed for electron microscopic observations. C, cis-Golgi network; G, Golgi apparatus; T, trans-Golgi network. Bars, 1 μm. (B) Size distribution of proacrosomal vesicles observed in SMAP2(+/+) (closed bars) and (−/−) (open bars) germ cells at the Golgi phase. Using the images shown in A, we measured diameters for each proacrosomal vesicle. The areas of the vesicles were calculated using image-processing software and divided by the square of the magnification. The diameters are presented in relative units. Vesicle sizes are binned as indicated. See Supplemental Figure S3 for the detailed size distribution of vesicles.

Mentions: Acrosome formation is initiated by the budding of proacrosomal vesicles from the TGN. To examine possible abnormalities in this step, we processed testes sections for analysis by transmission electron microscopy (TEM; Figure 5A). In the Golgi phase of acrosome formation, wild-type spermatids (Figure 5A, I) possess an umbrella-shaped TGN, and multiple proacrosomal vesicles of uniform size are present between the TGN and nuclear membrane. By contrast, in SMAP2-targeted spermatids (Figure 5A, II), proacrosomal vesicles were present but were not uniform in size and appeared to be larger than those in wild-type cells. In other SMAP2-targeted spermatids (Figure 5A, III), the lamellar structure of the TGN formed loose whorls. These observations suggest that there are abnormalities in the budding of vesicles from the TGN in SMAP2-targeted cells.


The Arf GAP SMAP2 is necessary for organized vesicle budding from the trans-Golgi network and subsequent acrosome formation in spermiogenesis.

Funaki T, Kon S, Tanabe K, Natsume W, Sato S, Shimizu T, Yoshida N, Wong WF, Ogura A, Ogawa T, Inoue K, Ogonuki N, Miki H, Mochida K, Endoh K, Yomogida K, Fukumoto M, Horai R, Iwakura Y, Ito C, Toshimori K, Watanabe T, Satake M - Mol. Biol. Cell (2013)

Electron microscopic observations of proacrosomal vesicle formation at the Golgi phase. (A) Germ cells from SMAP2(+/+) and (−/−) mice were processed for electron microscopic observations. C, cis-Golgi network; G, Golgi apparatus; T, trans-Golgi network. Bars, 1 μm. (B) Size distribution of proacrosomal vesicles observed in SMAP2(+/+) (closed bars) and (−/−) (open bars) germ cells at the Golgi phase. Using the images shown in A, we measured diameters for each proacrosomal vesicle. The areas of the vesicles were calculated using image-processing software and divided by the square of the magnification. The diameters are presented in relative units. Vesicle sizes are binned as indicated. See Supplemental Figure S3 for the detailed size distribution of vesicles.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3756916&req=5

Figure 5: Electron microscopic observations of proacrosomal vesicle formation at the Golgi phase. (A) Germ cells from SMAP2(+/+) and (−/−) mice were processed for electron microscopic observations. C, cis-Golgi network; G, Golgi apparatus; T, trans-Golgi network. Bars, 1 μm. (B) Size distribution of proacrosomal vesicles observed in SMAP2(+/+) (closed bars) and (−/−) (open bars) germ cells at the Golgi phase. Using the images shown in A, we measured diameters for each proacrosomal vesicle. The areas of the vesicles were calculated using image-processing software and divided by the square of the magnification. The diameters are presented in relative units. Vesicle sizes are binned as indicated. See Supplemental Figure S3 for the detailed size distribution of vesicles.
Mentions: Acrosome formation is initiated by the budding of proacrosomal vesicles from the TGN. To examine possible abnormalities in this step, we processed testes sections for analysis by transmission electron microscopy (TEM; Figure 5A). In the Golgi phase of acrosome formation, wild-type spermatids (Figure 5A, I) possess an umbrella-shaped TGN, and multiple proacrosomal vesicles of uniform size are present between the TGN and nuclear membrane. By contrast, in SMAP2-targeted spermatids (Figure 5A, II), proacrosomal vesicles were present but were not uniform in size and appeared to be larger than those in wild-type cells. In other SMAP2-targeted spermatids (Figure 5A, III), the lamellar structure of the TGN formed loose whorls. These observations suggest that there are abnormalities in the budding of vesicles from the TGN in SMAP2-targeted cells.

Bottom Line: In the present study, SMAP2 is detected on the TGN in the pachytene spermatocyte to the round spermatid stages of spermatogenesis.Furthermore, syntaxin2, a component of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex, is not properly concentrated at the site of acrosome formation.Thus this study reveals a link between SMAP2 and CALM/syntaxin2 in clathrin-coated vesicle formation from the TGN and subsequent acrosome formation.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Immunology, Department of Pathology, Institute of Development, Aging and Cancer, Tohoku University, Sendai 980-8575, Japan.

ABSTRACT
The trans-Golgi network (TGN) functions as a hub organelle in the exocytosis of clathrin-coated membrane vesicles, and SMAP2 is an Arf GTPase-activating protein that binds to both clathrin and the clathrin assembly protein (CALM). In the present study, SMAP2 is detected on the TGN in the pachytene spermatocyte to the round spermatid stages of spermatogenesis. Gene targeting reveals that SMAP2-deficient male mice are healthy and survive to adulthood but are infertile and exhibit globozoospermia. In SMAP2-deficient spermatids, the diameter of proacrosomal vesicles budding from TGN increases, TGN structures are distorted, acrosome formation is severely impaired, and reorganization of the nucleus does not proceed properly. CALM functions to regulate vesicle sizes, and this study shows that CALM is not recruited to the TGN in the absence of SMAP2. Furthermore, syntaxin2, a component of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex, is not properly concentrated at the site of acrosome formation. Thus this study reveals a link between SMAP2 and CALM/syntaxin2 in clathrin-coated vesicle formation from the TGN and subsequent acrosome formation. SMAP2-deficient mice provide a model for globozoospermia in humans.

Show MeSH
Related in: MedlinePlus