Analysis of the roles of RGD-binding integrins, α4/α9 integrins, α6 integrins, and CD9 in the interaction of the fertilin β (ADAM2) disintegrin domain with the mouse egg membrane

Fertilin β (also known as ADAM2), a mammalian sperm protein that mediates gamete cell adhesion during fertilization, is a member of the ADAM protein family whose members have disintegrin domains with homology to integrin ligands found in snake venoms. Fertilin β utilizes an ECD sequence within its disintegrin domain to interact with the egg plasma membrane; the Asp is especially critical. Based on what is known about different integrin subfamilies and their ligands, we sought to characterize fertilin β binding sites on mouse eggs, focusing on integrin subfamilies that recognize short peptide sequences that include an Asp residue: the α₅/α₈/α_v /α_IIb or RGD-binding subfamily α₅β₁, α₈β₁, α_vβ₁, α_vβ₃, α_vβ₅, α_vβ₆, α_vβ₈, and α_IIbβ₃) and the α₄/α₉ subfamily (α₄β₁, α₉β₁, and α₄β₇). We tested peptide sequences known to perturb interactions mediated by these integrins in two different assays for fertilin β binding. Peptides with the sequence MLDG, which perturb α₄/α₉ integrin-mediated interactions, significantly inhibit fertilin β binding to eggs, which suggests a role for a member of this integrin subfamily as a fertilin β receptor. RGD peptides, which perturb α₅/α₈/α_v/ α_IIb integrin-mediated interactions, have partial inhibitory activity. The anti-α₆ antibody GoH3 has little or no inhibitory activity. An antibody to the integrin-associated tetraspanin protein CD9 inhibits the binding of a multivalent presentation of fertilin β (immobilized on beads) but not soluble fertilin β, which we speculate has implications for the role of CD9 in the strengthening of fertilin β-mediated cell adhesion but not in initial ligand binding.