Post-Translational Modifications of Nuclear and Non-Nuclear Proteins in Spermatozoa

Posttranslational modifications of nuclear and nonnuclear proteins in spermatozoa Summary The number of couples who need the help of assisted reproductive technology (ART) has increased over the years. Approximately half of the cases are caused by male infertility, which is often diagnosed as idiopathic infertility. Therefore, the search for male fertility markers will improve male infertility diagnosis, thereby facilitating advanced sperm treatment and selection via ART. Posttranslational modifications (PTMs) of sperm nuclear and nonnuclear proteins are suitable candidates for such markers. The PTMs of protamines and histones reflect sperm chromatin maturity and its readiness for fertilization, and accordingly, they can predict the outcome of ART. However, the PTMs of nonnuclear proteins, including cytoplasmic, cytoskeletal, and membrane proteins, reflect the ability of sperm to undergo hyperactivation, capacitation, or acrosome reactions, which are processes essential for fertilization. We hypothesize that the PTMs of nuclear and nonnuclear proteins can reflect sperm quality and, thus, serve as a valuable marker in ART. Additionally, we suggest that the in vitro addition of hydrogen sulfide into the sperm-manipulating media improves sperm motility and viability via persulfidation. We used Western blot..

Post-translační modifikace jaderných a ne-jaderných proteinů ve spermii

Posttranslational modifications of nuclear and nonnuclear proteins in spermatozoa Summary The number of couples who need the help of assisted reproductive technology (ART) has increased over the years. Approximately half of the cases are caused by male infertility, which is often diagnosed as idiopathic infertility. Therefore, the search for male fertility markers will improve male infertility diagnosis, thereby facilitating advanced sperm treatment and selection via ART. Posttranslational modifications (PTMs) of sperm nuclear and nonnuclear proteins are suitable candidates for such markers. The PTMs of protamines and histones reflect sperm chromatin maturity and its readiness for fertilization, and accordingly, they can predict the outcome of ART. However, the PTMs of nonnuclear proteins, including cytoplasmic, cytoskeletal, and membrane proteins, reflect the ability of sperm to undergo hyperactivation, capacitation, or acrosome reactions, which are processes essential for fertilization. We hypothesize that the PTMs of nuclear and nonnuclear proteins can reflect sperm quality and, thus, serve as a valuable marker in ART. Additionally, we suggest that the in vitro addition of hydrogen sulfide into the sperm-manipulating media improves sperm motility and viability via persulfidation. We used Western blot...Posttranslační modifikace jaderných a nejaderných proteinů ve spermiích Souhrn Kvůli početí potomka vyhledává pomoc asistovaných reprodukčních technologií (ART) stále více párů. Až za polovinou případů párové neplodnosti, stojí mužská neplodnost, která nezřídka končí diagnózou idiopatická (bez známé příčiny). Hledání nových ukazatelů mužské plodnosti, má proto velký význam pro diagnostiku mužské neplodnisti a případně pro selekci spermiích (ne)vhodných pro použití v ART. Post-translační modifikace (PTM) jaderných a nejaderných proteinů se zdají být vhodnými kandidáty pro takové ukazatele. Na jedné straně jsou PTM jaderných proteinů, protamínů a histonů, které odrážejí zralost chromatinu spermie, jeho připravenost na oplození a také kvalitu budoucího embrya. Na druhé straně jsou PTM nejaderných proteinů, zahrnující cytoplasmatické, cytoskeletární a membránové proteiny, které odrážejí schopnost spermie podstoupit hyperaktivaci, kapacitaci a akrosomální reakci, jinými slovy procesy, které jsou nezbytné pro nalezení a oplození oocytu. V této práci předpokládáme, že PTM jaderných a nejaderných proteinů odrážejí kvalitu spermií, a tak mohou sloužit jako selektivní ukazatele pro využití v ART. Dále předpokládáme, že přidání hydrogen sulfanu (H2S) do in vitro manipulačního média, zlepší motilitu a viabilitu...Ústav histologie a embryologieLékařská fakulta v PlzniFaculty of Medicine in Pilse

Male SIRT1 insufficiency leads to sperm with decreased ability to hyperactivate and fertilize

Deficient sperm motility is a frequent cause of the age-related male sub-/infertility. Since the protein sirtuin 1 (SIRT1) develops anti-aging action and participates in sperm motility and ATP synthesis in mitochondria, we investigated its role in the acquisition of hyperactivated motility during capacitation. For this, the dynamics of sperm subpopulations were studied, using males of Sirt1+/− heterozygous mutant mice. After 2 hr of capacitation, we observed reduced percentage of hyperactivated spermatozoa in Sirt1+/− males. Interestingly, prior to capacitation, Sirt1+/− spermatozoa showed higher mitochondrial superoxide levels, which could render mitochondrial injury and thereby motility defects. Accordingly, the fertilization rate of Sirt1+/− males after mating was decreased. We elucidated that SIRT1 male insufficiency underlies posterior sperm defects to hyperactivate during capacitation and propose Sirt1+/− males as a model for the study of the age-related infertility.This study was supported by the Charles University Research Fund (Progress Q39) and by the Ministry of Education, Youth and Sports of the Czech Republic, due to project No. CZ.02.1.01/0.0/0.0/16_019/0000787 ‘Fighting Infectious Diseases’, and the grant SVV–2020-2022 No 260 536.Peer reviewe

Low Doses of Bisphenol S Affect Post-translational Modifications of Sperm Proteins in Male Mice

Background: Bisphenol S (BPS) is increasingly used as a replacement for bisphenol A in the manufacture of products containing polycarbonates and epoxy resins. However, further studies of BPS exposure are needed for the assessment of health risks to humans. In this study we assessed the potential harmfulness of low-dose BPS on reproduction in male mice. Methods: To simulate human exposure under experimental conditions, 8-week-old outbred ICR male mice received 8 weeks of drinking water containing a broad range of BPS doses [0.001, 1.0, or 100 µg/kg body weight (bw)/day, BPS1-3] or vehicle control. Mice were sacrificed and testicular tissue taken for histological analysis and protein identification by nano-liquid chromatography/mass spectrometry (MS) and sperm collected for immunodetection of acetylated lysine and phosphorylated tyrosine followed by protein characterisation using matrix-assisted laser desorption ionisation time-of-flight MS (MALDI-TOF MS). Results: The results indicate that compared to vehicle, 100 µg/kg/day exposure (BPS3) leads to 1) significant histopathology in testicular tissue; and, 2) higher levels of the histone protein γH2AX, a reliable marker of DNA damage. There were fewer mature spermatozoa in the germ layer in the experimental group treated with 1 µg/kg bw (BPS2). Finally, Western blot and MALDI-TOF MS studies showed significant alterations in the sperm acetylome and phosphorylome in mice treated with the lowest (BPS1) exposure (0.001 µg/kg/day). Conclusions: In summary, this range of qualitative and quantitative findings in young male mice raise the possibility that relatively low doses of BPS may impair mammalian reproduction through epigenetic modifications of sperm proteins

Low doses of Bisphenol S affect post-translational modifications of sperm proteins in male mice

[Background]: Bisphenol S (BPS) is increasingly used as a replacement for bisphenol A in the manufacture of products containing polycarbonates and epoxy resins. However, further studies of BPS exposure are needed for the assessment of health risks to humans. In this study we assessed the potential harmfulness of low-dose BPS on reproduction in male mice.[Methods]: To simulate human exposure under experimental conditions, 8-week-old outbred ICR male mice received 8 weeks of drinking water containing a broad range of BPS doses [0.001, 1.0, or 100 μg/kg body weight (bw)/day, BPS1–3] or vehicle control. Mice were sacrificed and testicular tissue taken for histological analysis and protein identification by nano-liquid chromatography/mass spectrometry (MS) and sperm collected for immunodetection of acetylated lysine and phosphorylated tyrosine followed by protein characterisation using matrix-assisted laser desorption ionisation time-of-flight MS (MALDI-TOF MS).[Results]: The results indicate that compared to vehicle, 100 μg/kg/day exposure (BPS3) leads to 1) significant histopathology in testicular tissue; and, 2) higher levels of the histone protein γH2AX, a reliable marker of DNA damage. There were fewer mature spermatozoa in the germ layer in the experimental group treated with 1 μg/kg bw (BPS2). Finally, western blot and MALDI-TOF MS studies showed significant alterations in the sperm acetylome and phosphorylome in mice treated with the lowest exposure (0.001 μg/kg/day; BPS1), although the dose is several times lower than what has been published so far.[Conclusions]: In summary, this range of qualitative and quantitative findings in young male mice raise the possibility that very low doses of BPS may impair mammalian reproduction through epigenetic modifications of sperm proteins.This study was supported by the Czech Health Research Council (grant no. NV18–01-00544); the Charles University Research Fund (Progres Q39); the National Sustainability Programme I (NPU I) Nr. LO1503 provided by the Ministry of Education, Youth and Sports of the Czech Republic (MEYS CR); project no. SVV 02690 awarded by MEYS CR; and project no. CZ.02.1.01/0.0/0.0/16_019/0000787 “Fighting Infectious Diseases“, awarded by MEYS CR and financed by The European Regional Development Fund, European Human Biomonitoring Initiative HBM4EU provided by H2020, and the United States Fulbright Commission (P001496 grant to P.R.M.).Peer reviewe