Seminal proteoforms from bulls with contrasting semen freezability: a story deciphered by top-down mass spectrometry

in Reproduction
Authors:
Arabela Guedes de Azevedo Viana Department of General Biology, Federal University of Viçosa, Viçosa, Brazil
The Scripps Research Institute, La Jolla, California, USA

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Fabio Pereira Gomes Virginia Commonwealth University, Richmond, Virginia, USA

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Abdullah Kaya Selcuk University, Konya, Turkey
Alta Genetics Inc., Groningen, Netherlands

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Jolene Diedrich The Scripps Research Institute, La Jolla, California, USA

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Einko Topper Alta Genetics Inc., Groningen, Netherlands

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Dario Di Silvestre National Research Council of Italy, Proteomics and Metabolomics Unit, Institute for Biomedical Technologies, Milan, Italy

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Mariana Machado-Neves Department of General Biology, Federal University of Viçosa, Viçosa, Brazil

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Erdogan Memili Cooperative Agricultural Research Center, College of Agriculture, Food and Natural Resources, Prairie View A&M University, Prairie View, Texas, USA

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Arlindo Alencar Moura Department of Animal Science, Federal University of Ceará, Fortaleza, Brazil

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John R Yates The Scripps Research Institute, La Jolla, California, USA

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Correspondence should be addressed to F P Gomes: pereiragomf@vcu.edu or to A A Moura: arlindo.moura@gmail.com or to J R Yates: jyates@scripps.edu

(A Guedes de Azevedo Viana and F P Gomes contributed equally to this work)

DOI:
https://doi.org/10.1530/REP-24-0051
Volume/Issue:
Volume 169: Issue 2
Article ID:
e240051
Article Type:
Research Article
Online Publication Date:
11 Jan 2025
Copyright:
© the author(s) 2025
Restricted access
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In brief

Bovine sperm and seminal plasma (SP) proteoform atlas was characterized using top-down proteomics. Specific post-translational modifications and protein truncations correlated with semen freezability, with potential links to sperm functional processes.

Abstract

Top-down proteomics was employed to construct a proteoform atlas of sperm and SP from bulls with low semen freezability (LF) and high semen freezability (HF). Sperm and seminal proteins were fractionated by tandem size exclusion chromatography (<30 kDa) and analyzed by reversed-phase liquid chromatography-tandem mass spectrometry. This approach enabled identification of 299 SP (from 46 families) and 267 sperm proteoforms (from 139 families). Seventy proteoforms belonging to beta-defensin 10, c-type natriuretic peptide (NPPC), caltrin, seminal ribonuclease, osteopontin and binder of sperm protein (BSP) 3 families were unique to HF bulls’ SP. LF seminal proteins had 77 unique proteoforms, including caltrin, NPPC, osteopontin, BSP3, serpin family A member 5 and β-NGF families. Proteoform families of SP in HF and LF bulls were related to Ca2+ uptake, capacitation, acrosome reaction, sperm protection, fertilization and proteolytic processes. Thirty-three proteoforms of NPPC, caltrin and cylicin-2 families were upregulated in HF sperm. Twenty-two proteoforms of caltrin, cylicin-2, adenosine triphosphate (ATP) synthases and malate dehydrogenase families were among those upregulated in LF sperm. Truncated and acetylated histone H2A and non-truncated and acetylated c-Myc-binding protein were prevalent in LF sperm. Cylicin-2 proteoforms were observed in HF and LF sperm, and truncated glyceraldehyde-3-phosphate dehydrogenases were observed only in HF sperm. In silico analyses indicated the enrichment of mitochondrial metabolic pathways in HF sperm, including fatty acid metabolism and TCA cycle. Our study provides an unprecedented description of the bovine SP and sperm proteoforms. Post-translational processing appears to define the bioproperties of semen proteins and their associations with sperm cryoresistance.

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Print ISSN:
1470-1626
Online ISSN:
1741-7899

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