Dataset Information

Characterization of CD46 and ?1 integrin dynamics during sperm acrosome reaction.

ABSTRACT: The acrosome reaction (AR) is a process of membrane fusion and lytic enzyme release, which enables sperm to penetrate the egg surroundings. It is widely recognized that specific sperm proteins form an active network prior to fertilization, and their dynamic relocation is crucial for the sperm-egg fusion. The unique presence of the membrane cofactor protein CD46 in the sperm acrosomal membrane was shown, however, its behaviour and connection with other sperm proteins has not been explored further. Using super resolution microscopy, we demonstrated a dynamic CD46 reorganisation over the sperm head during the AR, and its interaction with transmembrane protein integrins, which was confirmed by proximity ligation assay. Furthermore, we propose their joint involvement in actin network rearrangement. Moreover, CD46 and ?1 integrins with subunit ?3, but not ?6, are localized into the apical acrosome and are expected to be involved in signal transduction pathways directing the acrosome stability and essential protein network rearrangements prior to gamete fusion.

SUBMITTER: Frolikova M

PROVIDER: S-EPMC5036054 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature

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Characterization of CD46 and β1 integrin dynamics during sperm acrosome reaction.

Frolikova Michaela M Sebkova Natasa N Ded Lukas L Dvorakova-Hortova Katerina K

Scientific reports 20160926

The acrosome reaction (AR) is a process of membrane fusion and lytic enzyme release, which enables sperm to penetrate the egg surroundings. It is widely recognized that specific sperm proteins form an active network prior to fertilization, and their dynamic relocation is crucial for the sperm-egg fusion. The unique presence of the membrane cofactor protein CD46 in the sperm acrosomal membrane was shown, however, its behaviour and connection with other sperm proteins has not been explored further ...[more]

PMID: 27666019

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Project description:The male gamete is not completely mature after ejaculation and requires further events in the female genital tract to acquire fertilizing ability, including the processes of capacitation and acrosome reaction. In order to shed light on protein changes experienced by the sperm cell in preparation for fertilization, a comprehensive quantitative proteomic profiling based on isotopic peptide labeling and liquid chromatography followed by tandem mass spectrometry was performed on spermatozoa from three donors of proven fertility under three sequential conditions: purification with density gradient centrifugation, incubation with capacitation medium, and induction of acrosome reaction by exposure to the calcium ionophore A23187. After applying strict selection criteria for peptide quantification and for statistical analyses, 36 proteins with significant changes in their relative abundance within sperm protein extracts were detected. Moreover, the presence of peptide residues potentially harboring sites for post-translational modification was revealed, suggesting that protein modification may be an important mechanism in sperm maturation. In this regard, increased levels of proteins mainly involved in motility and signaling, both regulated by protein modifiers, were detected in sperm lysates following incubation with capacitation medium. In contrast, less abundant proteins in acrosome-reacted cell lysates did not contain potentially modifiable residues, suggesting the possibility that all those proteins might be relocated or released during the process. Protein-protein interaction analysis revealed a subset of proteins potentially involved in sperm maturation, including the proteins Erlin-2 (ERLIN2), Gamma-glutamyl hydrolase (GGH) and Transmembrane emp24 domain-containing protein 10 (TMED10). These results contribute to the current knowledge of the molecular basis of human fertilization. It should now be possible to further validate the potential role of the detected altered proteins as modulators of male infertility.

Project description:BackgroundIn rodents, the cell surface complement regulatory protein CD46 is expressed solely on the spermatozoal acrosome membrane. Ablation of the CD46 gene is associated with a faster acrosome reaction. Sperm from Apodemus flavicollis (yellow-necked field mice), A. microps (pygmy field mice) and A. sylvaticus (European wood mice) fail to express CD46 protein and exhibit a more rapid acrosome reaction rate than Mus (house mice) or BALB/c mice. A. agrarius (striped field mice) belong to a different Apodemus subgenus and have pronounced promiscuity and large relative testis size. The aim of this study was to determine whether A. agrarius sperm fail to express CD46 protein and, if so, whether A. agrarius have a faster acrosome reaction than Mus.MethodsReverse transcription polymerase chain reaction (RT-PCR) was used to assess whether A. agrarius transcribe testicular CD46 mRNA. RT-PCR was supplemented with 3'- and 5'-rapid amplification of cDNA ends to determine the complete nucleotide sequence of A. agrarius CD46. Fluorescence microscopy was used to assess whether CD46 protein is expressed by A. agrarius sperm. The acrosome status of A. agrarius sperm was calculated over time by immunocytochemistry using peanut agglutinin lectin.ResultsWe demonstrate that A. agrarius mice transcribe two unique alternatively spliced testicular CD46 mRNA transcripts, both lacking exon 7, which differ from those described previously in other Apodemus species. The larger A. agrarius CD46 transcript has an insert between exons 10 and 11 which, if translated, would result in a novel cytoplasmic tail. In addition, A. agrarius CD46 transcripts have an extended AU-rich 3'-untranslated region (UTR) and a truncated 5'-UTR, resulting in failure to express spermatozoal CD46 protein. We show that A. agrarius has a significantly faster spontaneous acrosome reaction rate than A. sylvaticus and Mus.ConclusionAbsence of CD46 protein expression is associated with acrosomal instability in rodents. A. agrarius mice express novel CD46 transcripts, resulting in the trade of spermatozoal CD46 protein expression for a rapid acrosome reaction rate, in common with other species of field mice. This provides a strategy to increase competitive sperm advantage for individuals, leading to faster fertilisation in this highly promiscuous genus.

Dataset Information

Characterization of CD46 and ?1 integrin dynamics during sperm acrosome reaction.

Publications

Characterization of CD46 and β1 integrin dynamics during sperm acrosome reaction.

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