Sequence-specific interaction between the disintegrin domain of mouse ADAM 3 and murine eggs: role of beta1 integrin-associated proteins CD9, CD81, and CD98.
ADAM 3 is a sperm surface glycoprotein that has been implicated in sperm-egg adhesion. Because little is known about the adhesive activity of ADAMs, we investigated the interaction of ADAM 3 disintegrin domains, made in bacteria and in insect cells, with murine eggs. Both recombinant proteins inhibited sperm-egg binding and fusion with potencies similar to that which we recently reported for the ADAM 2 disintegrin domain. Alanine scanning mutagenesis revealed a critical importance for the glutamine at position 7 of the disintegrin loop. Fluorescent beads coated with the ADAM 3 disintegrin domain bound to the egg surface. Bead binding was inhibited by an authentic, but not by a scrambled, peptide analog of the disintegrin loop. Bead binding was also inhibited by the function-blocking anti-alpha6 monoclonal antibody (mAb) GoH3, but not by a nonfunction blocking anti-alpha6 mAb, or by mAbs against either the alphav or beta3 integrin subunits. We also present evidence that in addition to the tetraspanin CD9, two other beta1-integrin-associated proteins, the tetraspanin CD81 as well as the single pass transmembrane protein CD98 are expressed on murine eggs. Antibodies to CD9 and CD98 inhibited in vitro fertilization and binding of the ADAM 3 disintegrin domain. Our findings are discussed in terms of the involvement of multiple sperm ADAMs and multiple egg beta1 integrin-associated proteins in sperm-egg binding and fusion. We propose that an egg surface "tetraspan web" facilitates fertilization and that it may do so by fostering ADAM-integrin interactions.
Takahashi
Y
Y
Department of Cell Biology, University of Virginia Health System, School of Medicine, Charlottesville, Virginia 22908, USA.
Bigler
D
D
Ito
Y
Y
White
J M
JM
D051722:ADAM Proteins; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism / Q000502:physiology*; D002352:Carrier Proteins / Q000378:metabolism / Q000502:physiology*; D002460:Cell Line; D019483:Disintegrins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D004331:Drosophila melanogaster; D005260:Female; D000072196:Fertilins; D027261:Fusion Regulatory Protein-1; D006801:Humans; D039503:Integrin alpha6; D019012:Integrin beta1 / Q000378:metabolism*; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D008565:Membrane Proteins; D008666:Metalloendopeptidases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D010063:Ovum / Q000378:metabolism / Q000502:physiology; D051381:Rats; D013084:Sperm-Ovum Interactions / Q000502:physiology*; D013329:Structure-Activity Relationship; D060225:Tetraspanin 28; D060245:Tetraspanin 29
D015703:Antigens, CD; C554798:CD81 protein, human; C554869:CD9 protein, human; D002352:Carrier Proteins; C554799:Cd81 protein, mouse; C554870:Cd9 protein, mouse; D019483:Disintegrins; D027261:Fusion Regulatory Protein-1; D039503:Integrin alpha6; D019012:Integrin beta1; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D060225:Tetraspanin 28; D060245:Tetraspanin 29; D051722:ADAM Proteins; C000606454:ADAM2 protein, human; C499566:ADAM3A protein, human; C000606458:Adam2 protein, mouse; C000606459:Adam2 protein, rat; C485429:Adam3 protein, mouse; D000072196:Fertilins; D008666:Metalloendopeptidases