Molecular Cloning and Subcellular Localization of Tektin2-Binding Protein 1 (Ccdc 172) in Rat Spermatozoa

Yamaguchi, Airi; Kaneko, Tadaaki; Inai, Tetsuichiro; Iida, Hiroshi

doi:10.1369/0022155413520607

Cited by 20 publications

(16 citation statements)

References 45 publications

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“…It functions in the maintenance of sperm structures and movement [35]. ODF could be consider as a marker of male infertility factor and defects of ODF can lead to abnormal morphology and infertility [36].…”

Section: Discussionmentioning

confidence: 99%

Proteomic identification of cryostress in epididymal spermatozoa

Yoon

Rahman

Kwon

et al. 2016

J Animal Sci Biotechnol

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Background: Cryopreservation of epididymal spermatozoa is important in cases in which it is not possible to collect semen using normal methods, as the sudden death of an animal or a catastrophic injury. However, the freezing and thawing processes cause stress to spermatozoa, including cold shock, osmotic damage, and ice crystal formation, thereby reducing sperm quality. We assessed the motility (%), motion kinematics, capacitation status, and viability of spermatozoa using computer-assisted sperm analysis and Hoechst 33258/chlortetracycline fluorescence staining. Moreover, we identified proteins associated with cryostress using a proteomic approach and performed western blotting to validate two-dimensional electrophoresis (2-DE) results using two commercial antibodies. Results: Cryopreservation reduced viability (%), motility (%), straight-line velocity (VSL), average path velocity (VAP), amplitude of lateral head displacement (ALH), and capacitated spermatozoa, whereas straightness (STR) and the acrosome reaction increased after cryopreservation (P < 0.05). Nine proteins were differentially expressed (two proteins decreased and seven increased) (>3 fold, P < 0.05) before and after cryopreservation. The proteins differentially expressed following cryopreservation are putatively related to several signaling pathways, including the ephrinR-actin pathway, the ROS metabolism pathway, actin cytoskeleton assembly, actin cytoskeleton regulation, and the guanylate cyclase pathway.

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Section: Discussionmentioning

confidence: 99%

Proteomic identification of cryostress in epididymal spermatozoa

Yoon

Rahman

Kwon

et al. 2016

J Animal Sci Biotechnol

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“…TEKT2-null sperm exhibited flagella bending and reduced motility []). Recently, Yamaguchi et al [54] identified a 36kDa TEKT2-binding protein1 associated with mitochondria of rat sperm flagella. It has been reported that Tektin 2 of hamster sperm becomes tyrosine phosphorylated during in vitro capacitation [55].…”

Section: Discussionmentioning

confidence: 99%

Identification and characterization of a bovine sperm acrosomal matrix protein and its mechanism of interaction with acrosomal hydrolases

et al. 2015

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Fertilization, the union of male and female gametes to create offspring, is an intricate biological process dependent upon several biochemical and physiological events. Our understanding of the functions of protein constituents of the outer acrosomal membrane-associated matrix complex (OMC) is limited. A highly purified OMC fraction isolated from bovine cauda sperm heads is comprised of 54, 50, 45, and 38–19kDa polypeptides. The objective of this study is to identify and to characterize the 45kDa (OMC45) polypeptide and to define its role in binding acrosomal hydrolases and to examine the fate of OMC45 polypeptide during the acrosome reaction. We isolated OMC45 polypeptide from the high-pH insoluble fraction of OMC. Proteomic analysis of OMC45 by MALDI–TOF–TOF yielded 8 peptides that matched the NCBI database sequence of Tektin 3 (TEKT3). Triton X–100–permeabilized cauda sperm exhibited intense staining of the acrosomal segment with anti–OMC45 and anti–TEKT3. The OMC45 polypeptide was solubilized by RIPA (radio-immunoprecipitation assay) buffer extraction. The solubilized fraction was subjected to immunoprecipitation analysis. The OMC45 polypeptide was recovered in the anti–OMC45 immunoprecipitation pellet. An identical blot stained with anti–TEKT3 exhibited the presence of TEKT3 polypeptide in the anti–OMC45 pellet. Our immunofluorescence and biochemical studies confirm the proteomics identification of OMC45 polypeptide; that it exhibits a sequence similarity to TEKT3. OMC45 glycoprotein possesses both N–linked and O–linked oligosaccharides. Deglycosylated OMC45 revealed a significant reduction in both acrosin and N–acetylglucosaminidase (NAGA) binding in comparison with acrosin and NAGA binding to a native OMC45 polypeptide, demonstrating the important role of oligosaccharides in hydrolase binding. OMC45 polypeptide is not released during the acrosome reaction but remains in the particulate cell subfraction, associated with the hybrid membrane complex.

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“…The expression levels of the ODF major components (ODF1, ODF2, ODF3, and ODF4) were generally low in abundance in asthenozoospermatic men (Zhao et al, 2018). Thus, ODFs are involved in maintaining spermatozoa structure and movement (Yamaguchi et al, 2014). The absence of ODFs results in non-functional tails and affects spermatozoa motility (Yuan, 2014).…”

Section: Effect Of Vitrification On Outer Dense Fibers and Vimentinmentioning

confidence: 99%

Unraveling Subcellular and Ultrastructural Changes During Vitrification of Human Spermatozoa: Effect of a Mitochondria-Targeted Antioxidant and a Permeable Cryoprotectant

Kumar

Wang

Isachenko

et al. 2021

Front. Cell Dev. Biol.

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Mitochondria-targeted antioxidants have great potential to counterbalance the generated reactive oxygen species (ROS) because they cross the inner membrane of the mitochondria. Still, their use was not reported in vitrified human spermatozoa. Our laboratory has successfully vitrified spermatozoa without the use of permeable cryoprotectants, but subcellular-level evidence was missing. Therefore, this study aimed to improve spermatozoa vitrification using a mitochondria-targeted antioxidant (mitoquinone, MitoQ), reveal ultrastructural changes in the spermatozoa due to the use of a permeable cryoprotectant, and report alterations of functional proteins during the spermatozoa vitrification process. For this, each of 20 swim-up-prepared ejaculates was divided into seven aliquots and diluted with a vitrification medium supplemented with varying concentrations of MitoQ (0.02 and 0.2 μM), glycerol (1, 4, and 6%), and a combination of MitoQ and glycerol. All aliquots were vitrified by the aseptic capillary method developed in our laboratory. The spermatozoa function assays revealed that the addition of either MitoQ (0.02 μM), glycerol (1%), or a combination of MitoQ (0.02 μM) and glycerol (1%) in the vitrification medium results in better or equivalent spermatozoa quality relative to the control. Transmission electron microscopy revealed that MitoQ protects the spermatozoa from undergoing ultrastructural alterations, but glycerol induced ultrastructural alterations during the vitrification process. Next, we performed label-free quantitative proteomics and identified 1,759 proteins, of which 69, 60, 90, and 81 were altered in the basal medium, 0.02 μM MitoQ, 1% glycerol, and Mito-glycerol groups, respectively. Actin, tubulins, and outer dense fiber proteins were not affected during the vitrification process. Some of the identified ubiquitinating enzymes were affected during spermatozoa vitrification. Only a few proteins responsible for phosphorylation were altered during vitrification. Similarly, several proteins involved in spermatozoa–egg fusion and fertilization (IZUMO1 and Tektin) were not affected during the vitrification process. In conclusion, MitoQ attenuates the vitrification-induced ultrastructural changes and alterations in the key proteins involved in spermatozoa functions and fertilization.

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Molecular Cloning and Subcellular Localization of Tektin2-Binding Protein 1 (Ccdc 172) in Rat Spermatozoa

Cited by 20 publications

References 45 publications

Proteomic identification of cryostress in epididymal spermatozoa

Proteomic identification of cryostress in epididymal spermatozoa

Identification and characterization of a bovine sperm acrosomal matrix protein and its mechanism of interaction with acrosomal hydrolases

Unraveling Subcellular and Ultrastructural Changes During Vitrification of Human Spermatozoa: Effect of a Mitochondria-Targeted Antioxidant and a Permeable Cryoprotectant

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