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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.


Progress of CD46 and β1 integrin relocation prior and during AR.(A) CD46 (green), PNA lectin (red). (B) β1 integrin (green), PNA lectin (red). The first column represents a schematic illustration of (A) CD46, (B) β1 integrin localization in the acrosome intact sperm (line I) and in sperm during the AR progress. (A) CD46 detection in intact acrosomal membranes (line I), and the residual outer acrosomal membrane (line II) (see the green arrow), the inner acrosomal membrane begins to emerge (see the red arrow). CD46 relocation progress during the AR is seen across the apical equatorial segment towards the whole equatorial segment and the whole sperm head. (B) β1 integrin is relocated across the apical equatorial segment towards the whole equatorial segment and the whole sperm head. In contrast to CD46, the residual acrosome cap and the apical equatorial segments were not detected. Scale bar represents 2 μm.
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f1: Progress of CD46 and β1 integrin relocation prior and during AR.(A) CD46 (green), PNA lectin (red). (B) β1 integrin (green), PNA lectin (red). The first column represents a schematic illustration of (A) CD46, (B) β1 integrin localization in the acrosome intact sperm (line I) and in sperm during the AR progress. (A) CD46 detection in intact acrosomal membranes (line I), and the residual outer acrosomal membrane (line II) (see the green arrow), the inner acrosomal membrane begins to emerge (see the red arrow). CD46 relocation progress during the AR is seen across the apical equatorial segment towards the whole equatorial segment and the whole sperm head. (B) β1 integrin is relocated across the apical equatorial segment towards the whole equatorial segment and the whole sperm head. In contrast to CD46, the residual acrosome cap and the apical equatorial segments were not detected. Scale bar represents 2 μm.

Mentions: Specific monoclonal antibodies (mAbs), see methods, were used to label the individual proteins and follow their localization over the sperm head. Epifluorescent microscopic observation detected distinct sperm head regions that showed a profound distribution of both proteins (Fig. 1) during the acrosome reaction, but not during the capacitation. However, the protein relocation depended on the length of capacitation, whether 60 or 90 minutes. When sperm were left to capacitate for 90 min, the protein relocation during acrosome reaction was faster compared to the group capacitated for only 60 min (Fig. 2). CD46 displayed five clearly distinct labelled regions, such as the acrosome cap, residual acrosome cap, apical equatorial segment, equatorial segment and the whole sperm head (Fig. 1A). For the β1 integrins, only three patterns could be followed, such as the acrosome cap, equatorial segment and whole sperm head (Fig. 1B, S1a,b).


Characterization of CD46 and β 1 integrin dynamics during sperm acrosome reaction
Progress of CD46 and β1 integrin relocation prior and during AR.(A) CD46 (green), PNA lectin (red). (B) β1 integrin (green), PNA lectin (red). The first column represents a schematic illustration of (A) CD46, (B) β1 integrin localization in the acrosome intact sperm (line I) and in sperm during the AR progress. (A) CD46 detection in intact acrosomal membranes (line I), and the residual outer acrosomal membrane (line II) (see the green arrow), the inner acrosomal membrane begins to emerge (see the red arrow). CD46 relocation progress during the AR is seen across the apical equatorial segment towards the whole equatorial segment and the whole sperm head. (B) β1 integrin is relocated across the apical equatorial segment towards the whole equatorial segment and the whole sperm head. In contrast to CD46, the residual acrosome cap and the apical equatorial segments were not detected. Scale bar represents 2 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f1: Progress of CD46 and β1 integrin relocation prior and during AR.(A) CD46 (green), PNA lectin (red). (B) β1 integrin (green), PNA lectin (red). The first column represents a schematic illustration of (A) CD46, (B) β1 integrin localization in the acrosome intact sperm (line I) and in sperm during the AR progress. (A) CD46 detection in intact acrosomal membranes (line I), and the residual outer acrosomal membrane (line II) (see the green arrow), the inner acrosomal membrane begins to emerge (see the red arrow). CD46 relocation progress during the AR is seen across the apical equatorial segment towards the whole equatorial segment and the whole sperm head. (B) β1 integrin is relocated across the apical equatorial segment towards the whole equatorial segment and the whole sperm head. In contrast to CD46, the residual acrosome cap and the apical equatorial segments were not detected. Scale bar represents 2 μm.
Mentions: Specific monoclonal antibodies (mAbs), see methods, were used to label the individual proteins and follow their localization over the sperm head. Epifluorescent microscopic observation detected distinct sperm head regions that showed a profound distribution of both proteins (Fig. 1) during the acrosome reaction, but not during the capacitation. However, the protein relocation depended on the length of capacitation, whether 60 or 90 minutes. When sperm were left to capacitate for 90 min, the protein relocation during acrosome reaction was faster compared to the group capacitated for only 60 min (Fig. 2). CD46 displayed five clearly distinct labelled regions, such as the acrosome cap, residual acrosome cap, apical equatorial segment, equatorial segment and the whole sperm head (Fig. 1A). For the β1 integrins, only three patterns could be followed, such as the acrosome cap, equatorial segment and whole sperm head (Fig. 1B, S1a,b).

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.