Dataset Information

Quantitative proteomics analysis of Styxl1-/- mouse sperm

ABSTRACT: In order to investigate the molecular mechanism of Styxl1 in the regulation of sperm flagella, sperm from Styxl1+/+ and Styxl1-/- mice were collected for quantitative proteomics analysis using TMT-based liquid chromatography-tandem mass spectrometry (LC-MS/MS) approach. A total of 6429 proteins were quantified, and 1517 proteins (p < 0.05, Foldchange > 2) were differentially expressed. Among these differential proteins, 986 proteins were downregulated (65%) and 531 proteins were upregulated (35%) in Styxl1-/- sperm.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Sperm

SUBMITTER: Yueshuai Guo

LAB HEAD: Xuejiang Guo

PROVIDER: PXD046203 | Pride | 2023-12-01

REPOSITORIES: Pride

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			Action	DRS
	Search.zip	Other
	TMT6_sperm_styxl1_HPRP_F10_20230123.raw	Raw
	TMT6_sperm_styxl1_HPRP_F11_20230123.raw	Raw
	TMT6_sperm_styxl1_HPRP_F12_20230123.raw	Raw
	TMT6_sperm_styxl1_HPRP_F13_20230124.raw	Raw

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Project description:Testosterone is essential to maintain qualitative spermatogenesis. Nonetheless, no studies have been yet performed in humans to analyze the testosterone-mediated expression of the sperm proteins and their importance in reproduction. Thus, this study aimed to identify sperm protein alterations in male hypogonadism using proteomic profiling. We have performed a comparative proteomic analysis comparing sperm from fertile controls (a pool of 5 normogonadic normozoospermic fertile men) versus sperm from patients with secondary hypogonadism (a pool of 5 oligozoospermic hypogonadic patients due to isolated LH deficiency). Sperm protein composition was analyzed, after peptide labelling with Isobaric Tags, via liquid chromatography followed by tandem mass spectrometry (LC-MS/MS) on an LTQ Velos-Orbitrap mass spectrometer. LC-MS/MS data were analyzed using Proteome Discoverer. Criteria used to accept protein identification included a false discovery rate (FDR) of 1% and at least 1 peptide match per protein. Up to 986 proteins were identified and, of those, 43 proteins were differentially expressed: 32 proteins were under-expressed and 11 were over-expressed in the pool of hypogonadic patients. Bioinformatic analyses were performed using UniProt Knowledgebase, and the Gene Ontology Consortium database based on PANTHER. Notably, 13 of these 43 differentially expressed proteins have been previously reported to be related to sperm function and spermatogenesis. Western blot analyses for A-Kinase Anchoring Protein 3 (AKAP3) and the Prolactin Inducible Protein (PIP) were used to confirm the proteomics data. In summary, a high-resolution mass spectrometry-based proteomic approach was used for the first time to describe alterations of the sperm proteome in secondary male hypogonadism. A panel of deregulated sperm proteins was identified, including Prosaposin, Ropporin-1B, SERPINA5, SMOC-1, SPANXB1, GSG1, ELSPBP1, fibronectin, 5-oxoprolinase, AKAP3, AKAP4, HYDIN, ROPN1B, ß-Microseminoprotein and Protein S100-A8, which might represent putative physiological in vivo targets for androgen deficiency.

Project description:Current human reproductive risk assessment methods rely on semen and serum hormone analyses, which are not easily comparable to the histopathological endpoints and mating studies used in animal testing. Because of these limitations, there is a need to develop universal evaluations that reliably reflect male reproductive function. We hypothesized that toxicant-induced testicular injury can be detected in sperm using mRNA transcripts as indicators of insult. To test this, we exposed adult male Fischer 344 rats to low doses of model testicular toxicants and classically characterized the testicular injury while simultaneously evaluating sperm mRNA transcripts from the same animals. Overall, this study aimed to: 1) identify sperm transcripts altered after exposure to the model testicular toxicant, 2,5-hexanedione (HD) using microarrays; 2) expand on the HD-induced transcript changes in a comprehensive time course experiment using qRT-PCR arrays; and 3) test these injury indicators after exposure to another model testicular toxicant, carbendazim (CBZ). Microarray analysis of HD-treated adult Fischer 344 rats identified 128 altered sperm mRNA transcripts when compared to control using linear models of microarray analysis (q < 0.05). All transcript alterations disappeared after 3 months of post-exposure recovery. In the time course experiment, time-dependent alterations were observed for 12 candidate transcripts selected from the microarray data based upon fold change and biological relevance, and 8 of these transcripts remained significantly altered after the 3-month recovery period (p < 0.05). In the last experiment, 8 candidate transcripts changed after exposure to CBZ (p < 0.05). The two testicular toxicants produced distinct molecular signatures with only 4 overlapping transcripts between them, each occurring in opposite directions. Overall, these results suggest that sperm mRNA transcripts are indicators of low dose toxicant-induced testicular injury in the rat. Rats were exposed to sub-chronic low doses of the Sertoli cell toxicant 2,5-hexanedione (HD) or water (control for HD) for 3 months. Some rats in each group underwent 3 months of post-exposure recovery.

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Quantitative proteomics analysis of Styxl1-/- mouse sperm

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