Comparative proteome analysis of changes in the 26S proteasome during oocyte maturation in goldfish

Abstract

Proteasomes are large, multi-subunit particles that act as the proteolytic machinery for most of the regulated intracellular protein degradation in eukaryotic cells. An alteration of proteasome function may be important for the regulation of the meiotic cell cycle. To study the change at the subunit level of the 26S proteasome during meiotic maturation, we purified 26S proteasomes from immature and mature oocytes of goldfish. Two-dimensional polyacrylamide gel electrophoresis was used to separate proteins. For differential analysis, whole spots of the 26S proteasome from goldfish oocytes were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry and database analysis. Four spots that were different (only detected in mature oocyte 265 proteasomes and not in immature ones) and four protein spots that were up- or down-regulated were identified unambiguously. The mature-specific spots were not 26S proteasome components but rather their interacting proteins, and were identified as chaperonin-containing TCP-1 subunits and myosin light chain. Minor spots of three subunits of the 20S core particle and one of the 19S regulatory particle showed meiotic cell cycle-dependent changes. These results demonstrate that modifications of proteasomal subunits and cell cycle phase-dependent interactions of proteins with proteasomes occur during oocyte maturation in goldfish.

Affinity Purification Strategy to Capture Human Endogenous Proteasome Complexes Diversity and to Identify Proteasome-interacting Proteins

“…Forty-eight proteins were found in common, 38 were specific to our work, and 44 were not identified by us. Among the common proteins, 36 subunits of the proteasome complexes or activator proteins could be observed as well as 11 other proteins (supplemental Data 4); some of these had already been reported as PIPs in yeast (25,28,47) or in other species (21) (Tables I and II). Among the 46 putative PIPs specifically identified by Huang and co-workers (32,46), six proteins are linked to the UPP, and 38 are not.…”

“…Seven proteins were already known as PIPs or are related to known PIPs like histones, actins, myosins, or 14-3-3 proteins (21,32). Others constitute new putative PIPs and will need further investigation to be confirmed.…”

“…Histones (32) or proteins of the actomyosin cytoskeleton signaling pathway (21,57) have already been observed in TAP purified proteasome preparations or co-immunoprecipitate with proteasomes. Subunit ␣ of the nascent polypeptide-associated complex (NAC) has never been reported as a PIP so far.…”

Proteomic analysis of porcine oocytes during in vitro maturation reveals essential role for the ubiquitin C-terminal hydrolase-L1

“…With the progress of technologies, including linear amplification of cDNA populations, it has been possible to consider gene profiling in this biological model of extremely limited availability (Robert et al 2001, Goto et al 2002, Dalbies-Tran & Mermillod 2003, Zeng & Schultz 2003. In addition, the number of functional proteomic analyses identifying biologically relevant candidate proteins which may be involved in the regulation of oocyte maturation, embryo development, or oviductal proteome is gradually increasing in spite of the limited availability of human germ cells and lack of complete genome sequences of other mammalian species (Georgiou et al 2005, Horiguchi et al 2006, Massicotte et al 2006, Vitale et al 2007.…”

Identifying Dynamic Interactors of Protein Complexes by Quantitative Mass Spectrometry

“…The CCT complex is a molecular chaperone that plays important roles in the folding of tubulin, actin, and other cytosolic proteins (34). Previously two subunits of goldfish CCT complex have been shown to interact with the 26 S proteasome directly (35). In mammalian cells, the CCT complex is suspected to be a substrate of the ubiquitin-proteasome pathway because CCT complex components accumulated after treatment with proteasome inhibitor (34).…”