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Characterization of CD46 and β 1 integrin dynamics during sperm acrosome reaction

View Article: PubMed Central - PubMed

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 β1 integrins with subunit α3, but not α6, 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.

No MeSH data available.


Related in: MedlinePlus

Dynamics of CD46 and β1/α6/α3 integrin captured by STED.(A) (line I) CD46 (green) is locked over the acrosome vesicle and β1 integrin (red) is confined to both the plasma and the outer acrosomal membrane with prominent labelling of the perforatorium (see the red arrow); (line II) the onset of AR with the membrane vesiculation is visible, as well as the visibility of CD46 and β1 integrin within the acrosome and plasma membranes (see green and red arrows); (line III) the loss of the acrosome including the CD46 and β1 integrin signal is visible. Relocation of CD46 and β1 integrin over the equatorial segment in patchy clusters can be recognised; (line IV) CD46 is localized through the inner acrosome membrane (see the green arrow), the equatorial segment and over the post-acrosome region. β1 integrin shows a similar pattern to CD46 and also remains localized in the perforatorium (see the red arrow). (B) (line I) α6 (green) and β1 (red) integrins occupy different regions of the intact sperm head, except the same localization in the plasma membrane over the acrosome and the hook. α6 integrin is continues to be spread over the equatorial segment, even prior to AR (see the green arrows). β1 integrin is further present in the outer acrosomal membrane (when compared with the CD46 dual staining in AI) and perforatorium (see the red arrows). (line II) The α3 (red) integrin pattern is clearly different to the α6 (green) subunit, but remarkably similar to the one of β1 (red, line I) in the acrosome region. α3 is expressed on in the outer acrosomal membrane and the plasma membrane over the acrosome. (line III) The differences in the localization of the α3 (red) integrin and CD46 (green) are visible. The α3 integrin subunit is detectable on the plasma (see also Fig. S3b for detail) and the outer acrosomal membrane, when CD46 is defined strictly to the acrosomal membranes only. DAPI (blue); Scale bar represents 1 μm.
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f5: Dynamics of CD46 and β1/α6/α3 integrin captured by STED.(A) (line I) CD46 (green) is locked over the acrosome vesicle and β1 integrin (red) is confined to both the plasma and the outer acrosomal membrane with prominent labelling of the perforatorium (see the red arrow); (line II) the onset of AR with the membrane vesiculation is visible, as well as the visibility of CD46 and β1 integrin within the acrosome and plasma membranes (see green and red arrows); (line III) the loss of the acrosome including the CD46 and β1 integrin signal is visible. Relocation of CD46 and β1 integrin over the equatorial segment in patchy clusters can be recognised; (line IV) CD46 is localized through the inner acrosome membrane (see the green arrow), the equatorial segment and over the post-acrosome region. β1 integrin shows a similar pattern to CD46 and also remains localized in the perforatorium (see the red arrow). (B) (line I) α6 (green) and β1 (red) integrins occupy different regions of the intact sperm head, except the same localization in the plasma membrane over the acrosome and the hook. α6 integrin is continues to be spread over the equatorial segment, even prior to AR (see the green arrows). β1 integrin is further present in the outer acrosomal membrane (when compared with the CD46 dual staining in AI) and perforatorium (see the red arrows). (line II) The α3 (red) integrin pattern is clearly different to the α6 (green) subunit, but remarkably similar to the one of β1 (red, line I) in the acrosome region. α3 is expressed on in the outer acrosomal membrane and the plasma membrane over the acrosome. (line III) The differences in the localization of the α3 (red) integrin and CD46 (green) are visible. The α3 integrin subunit is detectable on the plasma (see also Fig. S3b for detail) and the outer acrosomal membrane, when CD46 is defined strictly to the acrosomal membranes only. DAPI (blue); Scale bar represents 1 μm.

Mentions: A detailed mutual localization of CD46 and β1 integrins was visualised on freshly released epididymal sperm with an intact acrosome (Fig. 5AI, S2 video image). In this case, the presence of CD46 was detected solely in the acrosomal cap in both the outer and inner acrosomal membranes, but not on the plasma membrane covering the acrosome region. Contrary to that, the β1 integrin subunit was present in the plasma membrane where it was detected in the area of acrosome cap (AC) and the hook marking the shape of the apical and dorsal sperm head (Fig. 5AI; 5BI and Fig. 6III, see the arrows). Unlike CD46, β1 integrins are present in an intact sperm head only in the outer acrosomal membrane, but not in the inner one (Fig. 5AI) and only later during the AR is the protein relocated over the inner acrosome membrane, equatorial segment and eventually the whole sperm head.


Characterization of CD46 and β 1 integrin dynamics during sperm acrosome reaction
Dynamics of CD46 and β1/α6/α3 integrin captured by STED.(A) (line I) CD46 (green) is locked over the acrosome vesicle and β1 integrin (red) is confined to both the plasma and the outer acrosomal membrane with prominent labelling of the perforatorium (see the red arrow); (line II) the onset of AR with the membrane vesiculation is visible, as well as the visibility of CD46 and β1 integrin within the acrosome and plasma membranes (see green and red arrows); (line III) the loss of the acrosome including the CD46 and β1 integrin signal is visible. Relocation of CD46 and β1 integrin over the equatorial segment in patchy clusters can be recognised; (line IV) CD46 is localized through the inner acrosome membrane (see the green arrow), the equatorial segment and over the post-acrosome region. β1 integrin shows a similar pattern to CD46 and also remains localized in the perforatorium (see the red arrow). (B) (line I) α6 (green) and β1 (red) integrins occupy different regions of the intact sperm head, except the same localization in the plasma membrane over the acrosome and the hook. α6 integrin is continues to be spread over the equatorial segment, even prior to AR (see the green arrows). β1 integrin is further present in the outer acrosomal membrane (when compared with the CD46 dual staining in AI) and perforatorium (see the red arrows). (line II) The α3 (red) integrin pattern is clearly different to the α6 (green) subunit, but remarkably similar to the one of β1 (red, line I) in the acrosome region. α3 is expressed on in the outer acrosomal membrane and the plasma membrane over the acrosome. (line III) The differences in the localization of the α3 (red) integrin and CD46 (green) are visible. The α3 integrin subunit is detectable on the plasma (see also Fig. S3b for detail) and the outer acrosomal membrane, when CD46 is defined strictly to the acrosomal membranes only. DAPI (blue); Scale bar represents 1 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5036054&req=5

f5: Dynamics of CD46 and β1/α6/α3 integrin captured by STED.(A) (line I) CD46 (green) is locked over the acrosome vesicle and β1 integrin (red) is confined to both the plasma and the outer acrosomal membrane with prominent labelling of the perforatorium (see the red arrow); (line II) the onset of AR with the membrane vesiculation is visible, as well as the visibility of CD46 and β1 integrin within the acrosome and plasma membranes (see green and red arrows); (line III) the loss of the acrosome including the CD46 and β1 integrin signal is visible. Relocation of CD46 and β1 integrin over the equatorial segment in patchy clusters can be recognised; (line IV) CD46 is localized through the inner acrosome membrane (see the green arrow), the equatorial segment and over the post-acrosome region. β1 integrin shows a similar pattern to CD46 and also remains localized in the perforatorium (see the red arrow). (B) (line I) α6 (green) and β1 (red) integrins occupy different regions of the intact sperm head, except the same localization in the plasma membrane over the acrosome and the hook. α6 integrin is continues to be spread over the equatorial segment, even prior to AR (see the green arrows). β1 integrin is further present in the outer acrosomal membrane (when compared with the CD46 dual staining in AI) and perforatorium (see the red arrows). (line II) The α3 (red) integrin pattern is clearly different to the α6 (green) subunit, but remarkably similar to the one of β1 (red, line I) in the acrosome region. α3 is expressed on in the outer acrosomal membrane and the plasma membrane over the acrosome. (line III) The differences in the localization of the α3 (red) integrin and CD46 (green) are visible. The α3 integrin subunit is detectable on the plasma (see also Fig. S3b for detail) and the outer acrosomal membrane, when CD46 is defined strictly to the acrosomal membranes only. DAPI (blue); Scale bar represents 1 μm.
Mentions: A detailed mutual localization of CD46 and β1 integrins was visualised on freshly released epididymal sperm with an intact acrosome (Fig. 5AI, S2 video image). In this case, the presence of CD46 was detected solely in the acrosomal cap in both the outer and inner acrosomal membranes, but not on the plasma membrane covering the acrosome region. Contrary to that, the β1 integrin subunit was present in the plasma membrane where it was detected in the area of acrosome cap (AC) and the hook marking the shape of the apical and dorsal sperm head (Fig. 5AI; 5BI and Fig. 6III, see the arrows). Unlike CD46, β1 integrins are present in an intact sperm head only in the outer acrosomal membrane, but not in the inner one (Fig. 5AI) and only later during the AR is the protein relocated over the inner acrosome membrane, equatorial segment and eventually the whole sperm head.

View Article: PubMed Central - PubMed

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 β1 integrins with subunit α3, but not α6, 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.

No MeSH data available.


Related in: MedlinePlus