Dataset Information

Analysis of protein extracts enriched in cell surface proteins from Bos taurus unsexed and sexed (X and Y) spermatozoa.

ABSTRACT: The sex of the animals is of paramount importance in many animal production systems. This is particularly evident in the production of milk or in breeding programs focused on the production of female animals. In some cases, slaughter or euthanasia of animals of the unwanted sex becomes the only solution, highlighting ethical and economic concerns. As global demand for food continues to rise, the importance of addressing these issues becomes more evident. Reproductive technologies, such as sperm sexing techniques, may hold the key to addressing both animal welfare and the sustainability of animal production. The accessibility of cell surface proteins makes them promising targets for innovative sperm sexing techniques. Therefore, a comprehensive shotgun proteomic analysis was performed on protein lysates specifically enriched in cell surface proteins through biotinylation isolation obtained from Limousin bull unsexed semen (BU), Holsteins-Frisian sexed X-semen (BX; purity > 90%), and a pool of Angus and Asturian sexed Y-semen (BY; purity > 90%). According to this analysis, after removal of contaminants, a total of 149 were reliably quantified. BU samples had 18 proteins significantly upregulated and 1 protein (contaminant) significantly downregulated when comparing with BX samples, and 52 proteins significantly upregulated when comparing with BY samples. Regarding possible differences between the X- and Y-sperm proteome, it was found that 5 proteins were significantly upregulated in BX samples. After removal of contaminants, a total of 130 quantified proteins were further analyzed. According to the DeepTMHMM v.1.0.13 software, out of the 130 quantified proteins, 81 were predicted to be globular proteins. Additionally, 23 were predicted as alpha-helical transmembrane proteins, 9 having a signal peptide. A total of 26 proteins were solely predicted to have a signal peptide. Furthermore, through cross-referencing these results with Gene Ontology information obtained from UniProt and one-to-one fast orthology assignments using the eggNOG-mapper v2.1.7 tool, it was possible to identify 28 proteins that could potentially be accessible from the cell surface. Among these are 11 proteins that were identified in BX samples but not in BY samples, making them potential protein targets specific to X-chromosome-bearing spermatozoa. Out of the 11 proteins, 5 have the highest potential for application in sperm sexing techniques as they are predicted to be transmembrane proteins with at least one transmembrane region and were identified in 2 or 3 replicates of the BX samples. Moreover, one of them is significantly more expressed in BX samples than in BY samples. None of the proteins quantified in BY were exclusive of this condition. Moreover, a total of 1,614 deamidated peptides were identified, corresponding to 265 deamidation sites. Of these, 144 deamidated peptides were reliably quantified (3 valid intensity values in one of the experimental conditions), corresponding to 133 unique deamidation sites on 71 proteins. Additionally, 32 possible N-linked glycosylation sites were identified (NxS/T motifs).

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Bos Taurus (bovine)

SUBMITTER: Rui Vitorino

LAB HEAD: Bruno Colaço

PROVIDER: PXD048616 | Pride | 2025-05-06

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	H090487_2p_WNE25_PM2_PRC-5587_Colaco_Fardilha_2.raw	Raw
	H090489_2p_WNE25_PM2_PRC-5587_Colaco_Fardilha_5.raw	Raw
	H090491_2p_WNE25_PM2_PRC-5587_Colaco_Fardilha_8.raw	Raw
	H090493_2p_WNE25_PM2_PRC-5587_Colaco_Fardilha_11.raw	Raw
	H090495_2p_WNE25_PM2_PRC-5587_Colaco_Fardilha_3.raw	Raw

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Publications

The Surface Proteome of Bovine Unsexed and Sexed Spermatozoa.

Pinto-Pinho Patrícia P Quelhas Joana J Impens Francis F Dufour Sara S Van Haver Delphi D Lopes Graça G Rocha António A Pinto-Leite Rosário R Fardilha Margarida M Colaço Bruno B

Animals : an open access journal from MDPI 20250208 4

Current sperm sexing methods are costly and largely restricted to cattle, while immunological techniques targeting sex-specific membrane proteins may offer more economical alternatives. To advance these methods, understanding the proteomic differences between the cell membranes of X- and Y-chromosome-bearing spermatozoa is essential. This study aimed to characterize the cell surface proteome of bovine sperm and identify potential targets for sperm sexing through LC-MS/MS analysis. Cell surface p ...[more]

PMID: 40002966

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