Growing evidence of ubiquitin-proteasome system (UPS) involvement in boar sperm capacitation.

0482721 - BTÚ 2018 US eng C - Conference Paper (international conference)
Zigo, Michal - Kerns, K. - Šutovsky, M. - Šutovský, P.
Growing evidence of ubiquitin-proteasome system (UPS) involvement in boar sperm capacitation.
Programm and Abstract book of the 10th International Conference on Pig Reproduction. Columbia, Missouri: University of Missouri, 2017, s. 24-25.
[10th International Conference on Pig Reproduction. Columbia (US), 11.06.2017-14.06.2017]
R&D Projects: GA MŠMT(CZ) ED1.1.00/02.0109
Institutional support: RVO:86652036
Keywords : ubiquitin * proteasome * sperm capacitation
OECD category: Biochemical research methods

Ubiquitination is a stable, reversible posttranslational modification of target proteins by covalent ligation of the small chaperone protein ubiquitin, in a well characterized enzymatic cascade, with participation of ubiquitin activating, conjugating, ligating and deubiquitinating enzymes. Ubiquitination targets proteins for degradation/recycling by the 26S proteasome, a multi-subunit, ubiquitin-specific proteolytic holoenzyme. Participation of UPS in fertilization came to light only recently. Studies using human and non-human mammalian spermatozoa revealed the role of UPS in the regulation of multiple fertilization events, necessary for sperm fertilizing ability. The present, ongoing study investigates the changes in proteasome compartmentalization, subunit composition and activity during in vitro capacitation of fresh boar spermatozoa. Sequential treatments of ejaculated and capacitated spermatozoa under proteasome permissive/inhibiting conditions were used to isolate putative sperm proteasome-associated sperm proteins in a compartment-specific manner. Treated spermatozoa were screened with state of the art image-based flow cytometer to assess changes in acrosome integrity, and the quantity and electrophoretic band patterns of candidate UPS substrate-proteins and proteasomal subunits. Resultant protein fractions were screened by WB for posttranslational modifications of i) proteasomal subunits, and ii) candidate proteasomal substrate/interactor proteins that co-purify with sperm proteasomes. Differential proteomic approach employing 1D PAGE revealed differences in accumulated proteins at the molecular weights of 65, 49, 35 kDa. MS analysis revealed accumulation of proteins previously reported proteins as well as some novel proteins, such as acrosin, cathepsin F, alpha subunit of ATP synthase, and enzymes of citric acid cycle.
Permanent Link: http://hdl.handle.net/11104/0278123