issecting EPPIN protease inhibitor domains in sperm motility and fertilizing ability: repercussions for male contraceptive developmen

Author(s)

Silva, Alan A. S. ^[1] ; Raimundo, Tamiris R. F. ^[1] ; Mariani, Noemia A. P. ^[1] ; Kushima, Helio ^[1] ; Avellar, Maria Christina W. ^[2] ; Buffone, Mariano G. ^[3] ; Paula-Lopes, Fabiola F. ^[4] ; Moura, Marcelo T. ^[4] ; Silva, Erick J. R. ^[1]

Affiliation

^[1] Sao Paulo State Univ, Inst Biosci, Dept Biophys & Pharmacol, Rua Prof Dr Antonio Celso W Zanin S-N, BR-18618689 Botucatu, SP - Brazil

^[2] Univ Fed Sao Paulo, Dept Pharmacol, Escola Paulista Med, Sao Paulo, SP - Brazil

^[3] Consejo Nacl Invest Cient & Tecn, Inst Biol & Med Expt, Buenos Aires, DF - Argentina

^[4] Univ Fed Sao Paulo, Dept Biol Sci, Campus Diadema, Diadema, SP - Brazil

• Document type: Journal article
• Source: MOLECULAR HUMAN REPRODUCTION; v. 27, n. 12 DEC 2021.
• Web of Science Citations: 0

Abstract

EPPIN (epididymal protease inhibitor) is a mammalian conserved sperm-binding protein displaying an N-terminal WFDC (whey-acidic protein four-disulfide core) and a C-terminal Kunitz protease inhibitor domains. EPPIN plays a key role in regulating sperm motility after ejaculation via interaction with the seminal plasma protein SEMG1 (semenogelin-1). EPPIN ligands targeting the SEMG1 binding site in the Kunitz domain are under development as male contraceptive drugs. Nevertheless, the relative contributions of EPPIN WFDC and Kunitz domains to sperm function remain obscure. Here, we evaluated the effects of antibodies targeting specific epitopes in EPPIN's WFDC (Q20E antibody, Gln20-Glu39 epitope) and Kunitz (S21C and F21C antibodies, Ser103-Cys123 and Phe90-C110 epitopes, respectively) domains on mouse sperm motility and fertilizing ability. Computer-assisted sperm analysis showed that sperm co-incubation with S21C antibody (but not F21C antibody) lowered progressive and hyperactivated motilities and impaired kinematic parameters describing progressive (straight-line velocity; VSL, average path velocity; VAP and straightness; STR) and vigorous sperm movements (curvilinear velocity; VCL, amplitude of lateral head movement; ALH, and linearity; LIN) compared with control. Conversely, Q20E antibodyinduced milder inhibition of progressive motility and kinematic parameters (VAP, VCL and ALH). Sperm co-incubation with S21C or Q20E antibodies affected in vitro fertilization as revealed by reduced cleavage rates, albeit without changes in capacitation-induced tyrosine phosphorylation. In conclusion, we show that targeting specific epitopes in EPPIN Kunitz and WFDC domains inhibits sperm motility and capacitation-associated events, which decrease their fertilizing ability; nevertheless, similar observations in vivo remain to be demonstrated. Simultaneously targeting residues in S21C and Q20E epitopes is a promising approach for the rational design of EPPIN-based ligands with spermostatic activity. (AU)

• FAPESP's process: 19/13661-1 - Potential roles of EPPIN (Epididymal Protease Inhibitor) on post-testicular maturation of mouse spermatozoa: consequences for sperm function
• Grantee: Alan Andrew dos Santos Silva
• Support Opportunities: Scholarships in Brazil - Master
• FAPESP's process: 15/08227-0 - A study on EPPIN as a drug target for male contraception: developing a new animal model to test the efficacy of male contraceptive drugs
• Grantee: Erick José Ramo da Silva
• Support Opportunities: Research Grants - Young Investigators Grants
• FAPESP's process: 17/11363-8 - EPPIN interacting proteins on mouse sperm surface: identification, expression and potential physiological roles
• Grantee: Noemia Aparecida Partelli Mariani
• Support Opportunities: Scholarships in Brazil - Master