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Interactome Analysis of EZH2 Protein in Control and GSNO exposed Human Umbilical Vein Endothelial Cells

ABSTRACT: We wanted to explore the comprehensive interacting partners of EZH2 upon S-nitrosylation to understand the effect of such post-translational modifications. Mass spectrometric analysis revealed the comprehensive association map of EZH2 in control and GSNO-treated cells. Samples are labeled as CHIMS (immunoprecipitation with EZH2 antibody using untreated cells), GS100 (immunoprecipitation with EZH2 antibody using GSNO treated cells), CONTROL (immunoprecipitation with EZH2 antibody using untreated cells), GSNO GS100 (immunoprecipitation with EZH2 antibody using GSNO treated cells). These are two biological replicates from control and GSNO treated cells.

INSTRUMENT(S): Orbitrap Exploris 240

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Syamantak Majumder

LAB HEAD: Prof. Syamantak

PROVIDER: PXD050209 | Pride | 2025-03-13

REPOSITORIES: Pride

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Project description:Abstract Buffalo bulls are the backbone of Indian dairy industry, and the quality of semen donating bulls determine the overall production efficiency of buffalo dairy farms. Seminal plasma (SP) of bulls harbor millions of lipid bilayer nanovesicles known as extracellular vesicles (EVs). These EVs carry a heterogenous cargo of essential biomolecules including fertility associated proteins which contribute to the fertilizing potential of spermatozoa. In this study, we explored various parameters of EVs such as size, concentration and the complete proteome profiles of SP EVs from two distinct fertility groups in order to unravel the differentially abundant proteins that potentially affect the overall fertility of bulls. Through Dynamic Light Scattering (DLS) it was found that the purified EVs were present in 7 to 14 size exclusion chromatographic (SEC) fractions with sizes ranging from 145 to 256 nm in the high fertile (HF) and low fertile (LF) bulls. Nanoparticle Tracking Analysis (NTA) confirmed the size of seminal EVs up to 200 nm, and concentrations varying from 2.84 to 6.82 × 1011 and 3.57 to 7.74 × 1011 particles per ml in HF and LF bulls, respectively across all the fractions. No significant difference was observed in the size and concentration of seminal EVs between the HF and LF groups. We identified a total of 1862 and 1807 proteins in seminal EVs of HF and LF bulls, respectively using high throughput LC-MS/MS approach. Out of these total proteins, 1754 proteins were common in both the groups and about 87 proteins were highly abundant in HF group while 1292 were less abundant as compared to LF bulls. Gene ontology (GO) analysis, revealed that the highly abundant proteins in HF group were mainly part of the nucleus and involved in nucleosome assembly along with DNA binding. In addition, highly abundant proteins in EVs of HF group were found to be involved in spermatogenesis, motility, acrosome reaction, capacitation, gamete fusion, and cryotolerance. Two highly abundant proteins of HF EVs, namely protein disulfide-isomerase A4 (PDIA4) and gelsolin (GSN), were successfully immunolocalized on spermatozoa, indicating that these proteins might be transferred to spermatozoa through EVs. The proteins; PDIA4 and GSN are intricately associated with sperm-oocyte fusion and acrosome reaction, respectively, thus they are vital for regulating the fertilizing capacity of sperm. Our evidences clearly support that the protein repertoires in EVs and subsequently their presence on sperm, are strongly associated with sperm functions. Altogether, the current investigation clearly indicates that SPEVs possess crucial protein repertoires which are essential for enhancing the sperm fertilizing capacity.

			Action	DRS
	ASHIMA_DR_SYAMANTAK_BITS_PILANI_ANALYSIS_14122023.pdResult	Other
	CHIMS.pdResult	Other
	CHIMS.raw	Raw
	CONTROL.raw	Raw
	GS100_30mMs.pdResult	Other

Dataset Information

Interactome Analysis of EZH2 Protein in Control and GSNO exposed Human Umbilical Vein Endothelial Cells

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