Presence, processing, and localization of mouse ADAM15 during sperm maturation and the role of its disintegrin domain during sperm–egg binding

Abstract

Successful fertilization requires gametes to complete several stages, beginning with maturation and transport along the male and female reproductive tracts and ending with the interaction between the sperm and the egg. This last step involves sperm-egg adhesion and membrane fusion. ADAMs (disintegrin and metalloprotease domain proteins) are a family of membrane-anchored glycoproteins that are thought to play diverse roles in cell-cell adhesion through their interaction with integrins. This study analyzes the presence, location, processing, and possible role of ADAM15 in mouse sperm. The presence of ADAM15 in mouse spermatozoa was detected by Western blotting, which revealed that ADAM15 is post-translationally processed, during epididymal sperm maturation and the acrosome reaction. The 35 kDa antigen present in the acrosome-reacted sperm is the last proteolytic product of the 110/75 kDa ADAM15 found in noncapacitated sperm. This 35 kDa protein contains the disintegrin domain. By indirect immunofluorescence, ADAM15 was identified in the acrosomal region and along the flagellum of mouse spermatozoa. In acrosome-reacted sperm, ADAM15 was lost from the acrosomal region, but remained diffusely distributed throughout the head and flagellum. Furthermore, the ADAM15 disintegrin domain (RPPTDDCDLPEF) partially inhibited fusion and almost completely inhibited sperm-oolemma adhesion. In conclusion, our data indicate that ADAM15 is present in the testis and in spermatozoa from the caput, corpus, and cauda epididymis, as well as in non-capacitated and acrosome-reacted gametes. Results also indicate that ADAM15 is processed during epididymal maturation and acrosome reaction and that it may play a role during sperm-egg binding.

Calpain modulates capacitation and acrosome reaction through cleavage of the spectrin cytoskeleton

“…In the same way as the principal piece, the cytoskeleton associated with PM in the acrosomal region is mainly composed of spectrin/F-actin, but with the difference that it is remodeled during capacitation. During capacitation and AR, proteins associated with PM as well as with membrane lipids compartmentalize in the acrosome region; they are redistributed inside the acrosome or distribute towards the equatorial segment and postacrosomal region as well (Rochwerger & Cuasnicu 1992, Da Ros et al 2004, Selvaraj et al 2007, Tsai et al 2007, Pasten-Hidalgo et al 2008. These molecules in some way are maintained as compartmentalized in the acrosome before capacitation.…”

“…Capacitation in the presence and absence of ALLN or calpeptin Capacitation was performed as described elsewhere (CabelloAgueros et al 2003, Pasten-Hidalgo et al 2008; briefly, ductus deferens guinea pig sperm were obtained and washed in 154 mM NaCl solution. Sperm cells (3.5!10 7 cell/ml) were capacitated by incubation at 37 8C in MCM-PL without glucose.…”

“…Cells (350!10 6 ) were resuspended in lysis buffer (50 mM Tris-HCl, pH 7.4, 1 mM EGTA, 1 mM PMSF, Complete, 1 mg/ml aprotinin, 10 mM sodium orthovanadate, 25 mM sodium fluoride, and 1% Triton X-100) as previously reported (Pasten-Hidalgo et al 2008). Samples were then centrifuged at 5000 g for 5 min at 4 8C, supernatants were collected, and protein concentration was determined (Bradford 1976).…”

SPACA3gene variants in a New Zealand cohort of infertile and fertile couples

“…The major variant at this locus is conserved across all known SPACA3 orthologs, indicating its importance to the functioning of SPRASA. This variant is located within the seed region for hs-miR873, a miRNA Pasten-Hidalgo et al, 2008) that can specify posttranscriptional repression by transcript destabilisation/translational repression (Friedman et al, 2009). The identifi cation of hs-miR873 and the conservation of this region suggest that this locus may be important in the regulation of SPACA3 by affecting the stability and/or translation of the mRNA, thereby affecting levels of the SPRASA protein.…”

ADAM15 participates in fertilization through a physical interaction with acrogranin

“…The heterodimer ADAM1/ADAM2, also known as fertilin a/b, which plays an important role in sperm-egg interaction during fertilization, is the best-studied of the ADAM family (Myles et al 1994, Evans et al 1995, Cho et al 1998, Chen et al 1999. Another well-studied example is ADAM15, which is expressed on the surface of a wide range of somatic cells and in mouse spermatozoa, has a role as an adhesion protein (Herren et al 1997, Poghosyan et al 2002, Pasten-Hidalgo et al 2008. We have recently reported that ADAM15 is expressed in mouse spermatozoa.…”