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Proteasomal interference prevents zona pellucida penetration and fertilization in mammals.

Authors
Sutovsky-P; Manandhar-G; McCauley-TC; Caamano-JN; Sutovsky-M; Thompson-WE; Day-BN
Source
Biol Reprod 2004 Nov; 71(5):1625-1637
NIOSHTIC No.
20029411
Abstract
The ubiquitin-proteasome pathway has been implicated in the penetration of ascidian vitelline envelope by the fertilizing spermatozoon (Sawada et al., Proc Natl Acad Sci U S A 2002; 99:1223-1228). The present study provides experimental evidence demonstrating proteasome involvement in the penetration of mammalian zona pellucida (ZP). Using porcine in vitro fertilization as a model, penetration of ZP was completely inhibited by specific proteasomal inhibitors MG-132 and lactacystin. Three commercial rabbit sera recognizing 20S proteasomal core subunits beta-1i, beta-2i, alpha-6, and beta-5 completely blocked fertilization at a very low concentration (i.e., diluted 1/2000 to 1/8000 in fertilization medium). Neither proteasome inhibitors nor antibodies had any effects on sperm-ZP binding and acrosome exocytosis in zona-enclosed oocytes or on fertilization rates in zona-free oocytes, which were highly polyspermic. Consistent with a possible role of ubiquitin-proteasome pathway in ZP penetration, ubiquitin and various alpha and beta type proteasomal subunits were detected in boar sperm acrosome by specific antibodies, immunoprecipitated and microsequenced by MALDI-TOF from boar sperm extracts. Antiubiquitin-immunoreactive substrates were detected on the outer face of ZP by epifluorescence microscopy. This study therefore provides strong evidence implicating the ubiquitin-proteasome pathway in mammalian fertilization and zona penetration. This finding opens a new line of acrosome/ZP research because further studies of the sperm acrosomal proteasome can provide new tools for the management of polyspermia during in vitro fertilization and identify new targets for contraceptive development.
Keywords
Reproductive-system-disorders; Reproductive-system; Spermatogenesis; Spermatozoa; Fertility; Mammalian-cells; In-vitro-study; Immune-reaction; Immune-system; Mass-spectrometry; Microscopic-analysis
Contact
Department of Animal Science, University of Missouri-Columbia, S141 ASRC, 920 East Campus Dr., Columbia, MO 65211-5300
CODEN
BIREBV
Publication Date
20041101
Document Type
Journal Article
Email Address
sutovskyp@missouri.edu
Funding Amount
235845
Funding Type
Grant
Fiscal Year
2005
NTIS Accession No.
NTIS Price
Identifying No.
Grant-Number-R21-OH-007324
Issue of Publication
5
ISSN
0006-3363
Source Name
Biology of Reproduction
State
OR; MO
Performing Organization
University of Missouri, Columbia, Missouri
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