ADAM 20 and 21; two novel human testis-specific membrane metalloproteases with similarity to fertilin-α
Introduction
Recently a new family of proteins, called ADAMs, was identified whose members are type I integral membrane proteins characterized by a disintegrin and metalloprotease domain (see Blobel, 1997; Wolfsberg et al., 1995a, Wolfsberg et al., 1995b; Huovila et al., 1996for recent reviews). The ADAMs are related to the crotalid snake venom disintegrin metalloproteases (SVMPs) and matrix metalloprotease (MMP) families, whose members lack transmembrane domains. Full-length ADAM cDNAs all encode a signal peptide followed by proprotein, Zn2+-metalloprotease, disintegrin, transmembrane region and cytoplasmic tail. Members of this family have been found in many animal species including mammals, Xenopus (Alfandari et al., 1997), Drosophila (Fambrough et al., 1996; Rooke et al., 1996) and nematodes (Podbilewicz, 1996; GenBank H89394), but none has, as yet, been found in unicellular eukaryotes or plants. Nor are ADAM genes found in the sequenced genomes of bacteria or S. cerevisiae.
In spite of the ADAM's conservation of structural domains, they appear to play rather diverse roles in development.
ADAM 12 (meltrin-α) was shown to be involved in myoblast, and perhaps also osteoclast fusion (Yagami-Hiromasa et al., 1995).
Mammalian ADAM 10 (MADAM) can degrade myelin basic protein (Howard et al., 1996). Its insect homologue Kuzbanian is involved in axonal extension in Drosophila (Fambrough et al., 1996; Rooke et al., 1996). The proteolytic substrate for Kuzbanian, and perhaps also for mammalian ADAM 10 is the Notch receptor, which regulates neuron proliferation (see Blobel, 1997for a recent discussion and references).
The ADAM 11 encoding gene is rearranged in primary breast tumours (Emi et al., 1993).
Another ADAM, TACE (TNF-α convertase), is required for cleavage of TNF-α from its membrane-bound precursor (Black et al., 1997; Moss et al., 1997a, Moss et al., 1997b).
ADAM 1 and -2, also named fertilin-α/β or PH30-α/β are expressed as heterodimers on the posterior head of mammalian spermatocytes and play a role in oocyte adhesion and fusion (Wolfsberg et al., 1993; Myles et al., 1994; Carroll et al., 1995; Wolfsberg and White, 1996; Myles and Primakoff, 1997). In mouse, it has been shown that fertilin β binds the α6/β1 integrin on oocytes, and that this interaction is essential for sperm–egg binding (Almeida et al., 1995). This subunit encodes a non-functional metalloprotease domain, which is absent from the mature protein (Blobel et al., 1990). The α subunit encodes an active protease domain and encodes a putative fusion peptide postulated to be involved in cell–cell fusion (Blobel et al., 1992; White, 1992; Muga et al., 1994). Like fertilin β, fertilin α may also be directly involved in sperm–egg adhesion (Evans et al., 1997). Fertilins α and β have been found in rat, rabbit, mouse, macaque and two species of guinea pig. In humans only the β subunit has been cloned (Gupta et al., 1996; Vidaeus et al., 1997). Surprisingly, the only human fertilin α gene is non-functional (Jury et al., 1997). This leads to the question as to what gene(s), then, are involved in human sperm–egg fusion, since fertilin β lacks potential fusion peptides. The fertilins are of considerable interest because of their essential role in fertilization and application as contraceptive vaccine (Herr, 1996) or drug target. It has been shown that vaccination with fertilin results in complete infertility in a male animal model (Ramarao et al., 1996).
Here we report the cDNA cloning of two novel, closely related human ADAM members which are expressed exclusively in testis. Their encoded products show good sequence similarity and share other characteristics with fertilin-α from other species; it is possible that they functionally replace fertilin-α.
Section snippets
RT–PCR
OligodT-primed, ds cDNA was prepared (Marathon kit, ClonTech, Palo Alto, CA) from polyA-RNA isolated from tonsillar B-cells which had been purified by rosetting on sheep red blood cells. Ten nanograms of DNA was used as template in a PCR containing 30 mM Tricine (pH 8.4), 2 mM MgCl2, 5 mM β-mercaptoethanol, 0.1 mg/ml gelatin, 0.1% Thesit (Ponce and Micol, 1992), 0.2 mM of each of the four dNTPs (freshly prepared from buffered 100 mM solution; Pharmacia, Uppsala, Sweden), 0.3 pmol/μl of each
cDNA cloning of ADAM 20 and -21
The original aim of this study was to identify novel ADAMs that might be involved in shedding of adhesion molecules and cytokines, such as L-selectin, ICAM-3 and TNF-α. As there is evidence, and one concrete example (TACE; Black et al., 1997; Moss et al., 1997a) that disintegrin–metalloproteases, or ADAMs are involved in these processes, degenerate oligonucleotides were designed that encode the Zn2+-binding pocket (HEXGHNFG) consensus and conserved disintegrin domain (GEECDXG). An RT–PCR on
Discussion
We describe here the cDNA cloning of two novel, closely related members of the ADAM family of membrane metalloproteases, ADAM 20 and ADAM 21. Comparison of their predicted translation products with known ADAMs showed they are related to fertilin α, fertilin β and meltrin-γ (ADAM 1, 2 and 9). Meltrin-γ mRNA is ubiquitously expressed (Yagami-Hiromasa et al., 1995) and not particularly up-regulated in testis (data not shown). ADAM 20 and ADAM 21's exclusive expression in human testis and sequence
Acknowledgements
I thank Dr J.-F. Gauchat for B-cell cDNA and Dr J.-Y. Bonneyfoy for initiating this work and stimulating discussions. The ADAM 20 and ADAM 21 sequences have been desposited with GenBank under accession numbers AF029899 and AF029900.
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MMPs, ADAMs and ADAMTSs are associated with mammalian sperm fate
2023, TheriogenologyCryopreservation and egg yolk medium alter the proteome of ram spermatozoa
2018, Journal of ProteomicsCitation Excerpt :These proteins may have some role in capacitation due to their involvement in phosphorylation [66] and bicarbonate metabolism [67,68] respectively. In addition, two other proteins which were decreased after cryopreservation, ADAM20-like and β hexosaminidase (aka β-N-acetylglucosaminidase), are believed to play important roles in oocyte binding [69,70] and penetration [71]. While mechanisms of zona pellucida binding may be redundant, disruption of ADAMs results in total infertility of mice due to a lack of zona binding [72].
Phylogenetic and molecular evolution of the ADAM (A Disintegrin And Metalloprotease) gene family from Xenopus tropicalis, to Mus musculus, Rattus norvegicus, and Homo sapiens
2012, GeneCitation Excerpt :By now, multiple ADAMs have been identified and their functions have been determined. Among them, about half, namely ADAMs 2, 7, 18, 20, 21, 29, and 30, have been found to be expressed either exclusively or predominately in the testis of mammals (Cerretti et al., 1999; Choi et al., 2003; Hooft van Huijsduijnen, 1998; Wei et al., 2010). These testis-specific/predominate ADAMs are reported to be involved in spermatogenesis and sperm functions such as sperm–egg binding (ADAM 2) and sperm–egg fusion (ADAM20 and 21) (Hooft van Huijsduijnen, 1998) (Cho et al., 2000; Edwards et al., 2008; Han et al., 2009; Kim et al., 2006b; Nishimura et al., 2001; Stein et al., 2005).
The ADAMs family: Coordinators of nervous system development, plasticity and repair
2006, Progress in NeurobiologyDefending the Zygote: Search for the Ancestral Animal Block to Polyspermy
2005, Current Topics in Developmental BiologyCitation Excerpt :Initially, the low sequence similarity among fertilin orthologs within the fusion peptide domain and the nonfunctional human fertilin α pseudogene suggests that fertilin proper is likely not involved in fusion of all eutherian gametes (see Jury et al., 1997, 1998; Myles and Primakoff, 1997). The discovery of complete fertilin α paralogs in mice (Nishimura et al., 2002) and primates (Hooft van Huijsduijnen, 1998; Jury et al., 1998), however, indicates that alternate forms of the fertilin heterodimer can exist. These neighboring intron‐less genes encode fertilin α paralogs of different mass with one paralog displaying a carboxy‐terminal truncation while another in humans lacks an appropriate active catalytic histidine (Hooft van Huijsduijnen, 1998; Jury et al., 1998; Nishimura et al., 2002).