Calcium-calmodulin and pH regulate protein tyrosine phosphorylation in stallion sperm.
Abstract: The mechanisms leading to capacitation in stallion sperm are poorly understood. The objective of our study was to define factors associated with regulation of protein tyrosine phosphorylation in stallion sperm. Stallion sperm were incubated for 4 h in modified Whitten's media with or without bicarbonate, calcium, or BSA. When sperm were incubated in air at 30×10⁶/ml at initial pH 7.25, protein tyrosine phosphorylation was detected only in medium containing 25 mM bicarbonate alone; calcium and BSA inhibited phosphorylation. Surprisingly, this inhibition did not occur when sperm were incubated at 10×10⁶/ml. The final pH values after incubation at 30×10⁶ and 10×10⁶ sperm/ml were 7.43 ± 0.04 and 7.83 ± 0.07 (mean ± s.e.m.) respectively. Sperm were then incubated at initial pH values of 7.25, 7.90, or 8.50 in either air or 5% CO₂. Protein tyrosine phosphorylation increased with increasing final medium pH, regardless of the addition of bicarbonate or BSA. An increase in environmental pH was observed when raw semen was instilled into the uteri of estrous mares and retrieved after 30 min (from 7.47 ± 0.10 to 7.85 ± 0.08), demonstrating a potential physiological role for pH regulation of capacitation. Sperm incubated in the presence of the calmodulin (CaM) inhibitor W-7 exhibited a dose-dependent increase in protein tyrosine phosphorylation, suggesting that the inhibitory effect of calcium was CaM mediated. These results show for the first time a major regulatory role of external pH, calcium, and CaM in stallion sperm protein tyrosine phosphorylation.
Publication Date: 2012-07-27 PubMed ID: 22843772DOI: 10.1530/REP-12-0067Google Scholar: Lookup
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- Comparative Study
- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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The research article focuses on understanding the mechanisms leading to capacitation in stallion sperm by specifically examining factors regulating protein tyrosine phosphorylation, like pH and calcium-calmodulin. The findings show that external pH, calcium, and calcium’s interaction with a protein called calmodulin play a significant role.
Objective and Methodology
- The purpose of the study was to define factors related to the regulation of protein tyrosine phosphorylation in stallion sperm. The protein tyrosine phosphorylation is a process essential for various intercellular communications.
- Stallion sperm were incubated for 4 hours in an altered form of Whitten’s media (often used for testing sperm quality) with or without the presence of bicarbonate, calcium, or BSA (bovine serum albumin).
Results
- It was observed that protein tyrosine phosphorylation was detected only in the medium containing 25 mM bicarbonate alone when sperm were incubated at a very high concentration (30×10⁶/ml) and at an initial pH of 7.25. Both calcium and BSA inhibited phosphorylation under these conditions.
- However, this inhibitive effect was not seen when sperm were incubated at a lower concentration (10×10⁶/ml).
- They observed different final pH values based on sperm concentration, with higher pH noticed with lower concentration.
- An increase in protein tyrosine phosphorylation was noticed with increasing the final medium pH, regardless of the addition of bicarbonate or BSA, indicating that higher pH supports this process.
- When the calmodulin inhibitor (W-7) was present, there was a dose-dependent increase in protein tyrosine phosphorylation, suggesting that calcium’s inhibitory effect was mediated by its interaction with the protein calmodulin.
Significance
- This study, for the first time, demonstrates a significant regulatory role for factors like environmental pH, calcium, and calmodulin in the process of protein tyrosine phosphorylation in stallion sperm.
- These findings not only increase our understanding of stallion sperm capacitation but also show potential pathways that could be exploited for therapeutic measures or enhancing reproductive success in stallions.
Cite This Article
APA
González-Fernández L, Macías-García B, Velez IC, Varner DD, Hinrichs K.
(2012).
Calcium-calmodulin and pH regulate protein tyrosine phosphorylation in stallion sperm.
Reproduction, 144(4), 411-422.
https://doi.org/10.1530/REP-12-0067 Publication
Researcher Affiliations
- Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine & Biomedical Sciences, Texas A&M University, College Station, TX 77843-4466, USA.
MeSH Terms
- Animals
- Calcium / analysis
- Calcium Signaling / drug effects
- Calcium-Calmodulin-Dependent Protein Kinases / pharmacology
- Calmodulin / antagonists & inhibitors
- Calmodulin / metabolism
- Chelating Agents / pharmacology
- Egtazic Acid / pharmacology
- Enzyme Inhibitors
- Horses / physiology
- Hydrogen-Ion Concentration
- Male
- Phosphoproteins / metabolism
- Phosphorylation / drug effects
- Protein Processing, Post-Translational / drug effects
- Semen / chemistry
- Sperm Capacitation / drug effects
- Spermatozoa / drug effects
- Spermatozoa / metabolism
- Sulfonamides / pharmacology
- Tyrosine / metabolism
Citations
This article has been cited 6 times.- Felix MR, Turner RM, Dobbie T, Hinrichs K. Successful in vitro fertilization in the horse: production of blastocysts and birth of foals after prolonged sperm incubation for capacitation†.. Biol Reprod 2022 Dec 10;107(6):1551-1564.
- Martín-Hidalgo D, Macías-García B, González-Fernández L. Influence of different cellular concentrations of boar sperm suspensions on the induction of capacitation and acrosome reaction.. J Reprod Dev 2022 Feb 18;68(1):68-73.
- Bucci D, Spinaci M, Galeati G, Tamanini C. Different approaches for assessing sperm function.. Anim Reprod 2020 May 22;16(1):72-80.
- Touré A. Importance of SLC26 Transmembrane Anion Exchangers in Sperm Post-testicular Maturation and Fertilization Potential.. Front Cell Dev Biol 2019;7:230.
- Leemans B, Stout TAE, Soom AV, Gadella BM. pH-dependent effects of procaine on equine gamete activation†.. Biol Reprod 2019 Nov 21;101(5):1056-1074.
- González-Fernández L, Macías-García B, Calle-Guisado V, García-Marín LJ, Bragado MJ. Calmodulin inhibitors increase the affinity of Merocyanine 540 for boar sperm membrane under non-capacitating conditions.. J Reprod Dev 2018 Oct 12;64(5):445-449.
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