Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.25.1 (proteasome)
28,817 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Theophylline (THP) and 1,3-dinitrobenzene (DNB) are thought to induce infertility by incapacitating the nurturing Sertoli cells and causing germ cell apoptosis in the testicular seminiferous epithelium, respectively. We hypothesized that THP and DNB exposure would alter the expression of the genes within the ubiquitin-proteasome pathway (UPP), implicated in spermatogenesis and epididymal sperm quality control. Rats were fed 0 or 8000 ppm of THP and necropsied on Days 18, 30, and 42 or administered 0, 2, or 6 mg/kg DNB via oral gavage and necropsied on Day 7. Tissues were collected from the testis and the caput, corpus, and cauda regions of the epididymis for transcriptional profiling by semiquantitative RT-PCR, real-time RT-PCR, and histopathology. Target UPP genes included those encoding for constitutive the 20S proteasomal core subunits Psmb1 (beta1), Psmb2 (beta2), and Psmb5 (beta5); the inducible 20S core subunits Psmb9 (LMP2), Psmb8 (LMP7), and Psmb10 (LMP10); and Ube1 (ubiquitin-activating enzyme E1), Ube2d3 (ubiquitin-conjugating enzyme E2), and Uchl1 (ubiquitin C-terminal hydrolase PGP9.5). Spermatozoa were collected from the cauda epididymis for analysis by light microscopy and flow cytometric evaluation of sperm surface ubiquitin. These data show that reprotoxic exposure alters the tissue-specific expression of UPP genes in the testis and epididymis, which may contribute to the aberrant spermatogenesis and epididymal processing of both normal and defective spermatozoa. Transcriptional profiling and flow cytometric analysis of the UPP thus captures the prodromal effects of reproductive toxicity not captured by conventional histology and functional cytology. Complementing seminal analysis with these measures may be useful in screening drug-induced toxicity or environmental infertility.
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PMID:Differential expression of genes encoding constitutive and inducible 20S proteasomal core subunits in the testis and epididymis of theophylline- or 1,3-dinitrobenzene-exposed rats. 1698 15

mUBPy is a deubiquitinating enzyme expressed preferentially in male germ cells and neurons. Recently, mUBPy has been shown to be involved in the down-regulation of growth factor receptors. In mouse spermatozoa mUBPy interacts with the sperm-specific molecular chaperone MSJ-1 and associates with the proteasome. The ubiquitin/proteasome system plays a key role during spermatogenesis to yield functional spermatozoa. Immunoelectron microscopy has been here used to localize both mUBPy and MSJ-1 in mouse spermatozoa. mUBPy and MSJ-1 label the cytoplasmic side of the acrosomal membrane and the centrosome, two sperm structures fundamental for a successful fertilization. In vitro protein interaction assay reveals that mUBPy is able to bind gamma-tubulin, a centrosomal protein marker. This protein interaction has been confirmed in vivo by double protein immunolabelling in spermatogenic cells. Upon the grounds of these findings and in the light of recent acquisition on the centrosome biology, we suggest that mUBPy could have a key role during mouse fertilization and propose mUBPy as a novel centrosomal component.
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PMID:MUBPy is a novel centrosome-associated protein and interacts with gamma-tubulin. 1728 68

The ubiquitin-proteasome is an ubiquitous system mainly devoted to protein degradation. The presence of ubiquitinated proteins in male gametes suggests a role for this system also in reproduction. Available evidence indicate that ubiquitin in spermatozoa may have a role in semen quality control, as ubiquitinated defective spermatozoa in the epididymis are subsequently phagocytosed by epididymal epithelial cells. Moreover, a role both in the regulation of mitochondrial inheritance in mammals (paternal mitochondria are eliminated and their ubiquitination appears to be important for this process) and in sperm-oocyte interaction at fertilization (which is inhibited by an inhibitor of proteasome) have been also suggested. We found that both morphologically normal and abnormal human spermatozoa in semen may be ubiquitinated and that the percentage of ubiquitinated sperm in the ejaculate positively correlates with normal morphology and motility, suggesting that sperm ubiquitination may have a positive role in sperm functions. It remains to be defined if and which patterns of ubiquitination of spermatozoa may distinguish between the different biological functions of this system. In an attempt to answer this question, we set up a method to detect simultaneously ubiquitination and DNA fragmentation by FACScan since the latter parameter is related to a poor quality of semen; in particular, abnormal morphology. We found that DNA fragmented human spermatozoa are also ubiquitinated. Studies are in progress to determine the correlation between the fraction of ubiquitinated-non DNA fragmented spermatozoa and parameters of semen analysis.
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PMID:Biological meaning of ubiquitination and DNA fragmentation in human spermatozoa. 1756 70

We have shown that the proteasome is present in mammalian sperm and plays a role during fertilisation. In this work we studied the relationship between protein phosphorylation and proteasomal activity in human sperm. Aliquots of motile sperm were incubated for 0, 5 and 18 h at 37 degrees C, 5% CO2, with different concentration of the kinase inhibitors genistein, H89 or tamoxifen. Control aliquots were treated with the inhibitor solvent. The chymotrypsin-like activity of the proteasome was assayed using a fluorogenic substrate. Aliquots of spermatozoa capacitated during 18 h were incubated for 30 min with kinase inhibitors and then with 7 microM progesterone (P). The percentage of viable acrosome-reacted sperm was evaluated using FITC-labeled Pisum sativum agglutinin. The results indicate that spermatozoa treated with different concentrations of genistein and tamoxifen did not modify the chymotrypsin-like activity of the proteasome during capacitation. On the other hand, proteasome activity was significantly decreased by incubation with H89. Sperm treatment with genistein, H89 and tamoxifen significantly inhibited the P-induced acrosome reaction. Western blot analysis indicated that the proteasome inhibitor, epoxomicin, reduced serine protein phosphorylation. These results suggest that the enzymatic activity of the proteasome is modulated by protein kinase A, and that both enzymes are involved in the P-induced acrosome reaction.
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PMID:Proteasome activity and its relationship with protein phosphorylation during capacitation and acrosome reaction in human spermatozoa. 1764 68

The 26S proteasome, which is a multi-subunit protease with specificity for substrate proteins that are postranslationally modified by ubiquitination, has been implicated in acrosomal function and sperm-zona pellucida (ZP) penetration during mammalian fertilization. Ubiquitin C-terminal hydrolases (UCHs) are responsible for the removal of polyubiquitin chains during substrate priming for proteasomal proteolysis. The inhibition of deubiquitination increases the rate of proteasomal proteolysis. Consequently, we have hypothesized that inhibition of sperm acrosome-borne UCHs increases the rate of sperm-ZP penetration and polyspermy during porcine in vitro fertilization (IVF). Ubiquitin aldehyde (UA), which is a specific nonpermeating UCH inhibitor, significantly (P < 0.05) increased polyspermy during porcine IVF and reduced (P < 0.05) UCH enzymatic activity measured in motile boar spermatozoa using a specific fluorometric UCH substrate, ubiquitin-AMC. Antibodies against two closely related UCHs, UCHL1 and UCHL3, detected these UCHs in the oocyte cortex and on the sperm acrosome, respectively, and increased the rate of polyspermy during IVF, consistent with the UA-induced polyspermy surge. In the oocyte, UCHL3 was primarily associated with the meiotic spindle. Sperm-borne UCHL3 was localized to the acrosomal surface and coimmunoprecipitated with a peripheral acrosomal membrane protein, spermadhesin AQN1. Recombinant UCHs, UCHL3, and isopeptidase T reduced polyspermy when added to the fertilization medium. UCHL1 was detected in the oocyte cortex but not on the sperm surface, and was partially degraded 6-8 h after fertilization. Enucleated oocyte-somatic cell electrofusion caused polarized redistribution of cortical UCHL1. We conclude that sperm-acrosomal UCHs are involved in sperm-ZP interactions and antipolyspermy defense. Modulation of UCH activity could facilitate the management of polyspermy during IVF and provide insights into male infertility.
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PMID:Ubiquitin C-terminal hydrolase-activity is involved in sperm acrosomal function and anti-polyspermy defense during porcine fertilization. 1767 Dec 68

Spermatogenesis in mammals necessitates an extensive remodeling and loss of many cellular organelles and proteins as the spermatozoa undergo maturation. The removal of proteins and organelles depends on the ubiquitin-proteasome pathway. Here we show that the E3 ubiquitin ligase Herc4, though ubiquitously expressed in all tissues, is most highly expressed in the testis, specifically during spermiogenesis. Mice homozygous for a Herc4 mutation are overtly normal; however, overall the males produce litter sizes some 50% smaller whereas female homozygotes show normal fertility. The reduced fertility in males is associated with about 50% of mature spermatozoa having reduced motility. Many of the spermatozoa possess an angulated tail with a cytoplasmic droplet being retained at the angulation. Our results show that Herc4 ligase is required for proper maturation and removal of the cytoplasmic droplet for the spermatozoon to become fully functional.
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PMID:Disruption of the ubiquitin ligase HERC4 causes defects in spermatozoon maturation and impaired fertility. 1796 48

Posttranslational modification by ubiquitination marks defective or outlived intracellular proteins for proteolytic degradation by the 26S proteasome. The ATP-dependent, covalent ligation and formation of polyubiquitin chains on substrate proteins requires the presence and activity of a set of ubiquitin activating and conjugating enzymes. While protein ubiquitination typically occurs in the cell cytosol or nucleus, defective mammalian spermatozoa become ubiquitinated on their surface during post-testicular sperm maturation in the epididymis, suggesting an active molecular mechanism for sperm quality control. Consequently, we hypothesized that the bioactive constituents of ubiquitin-proteasome pathway were secreted in the mammalian epididymal fluid (EF) and capable of ubiquitinating extrinsic substrates. Western blotting indeed detected the presence of the ubiquitin-activating enzyme E1 and presumed E1-ubiquitin thiol-ester intermediates, ubiquitin-carrier enzyme E2 and presumed E2-ubiquitin thiol-ester intermediates and the ubiquitin C-terminal hydrolase PGP 9.5/UCHL1 in the isolated bovine EF. Thiol-ester assays utilizing recombinant ubiquitin-activating and ubiquitin-conjugating enzymes, biotinylated substrates, and isolated bovine EF confirmed the activity of the ubiquitin activating and conjugating enzymes within EF. Ubiquitinated proteins were found to be enriched in the defective bull sperm fraction and appropriate proteasomal deubiquitinating and proteolytic activities were measured in the isolated EF by specific fluorescent substrates. The apocrine secretion of cytosolic proteins was visualized in transgenic mice and rats expressing the enhanced green fluorescent protein (eGFP) under the direction of ubiquitin-C promoter. Accumulation of eGFP, ubiquitin and proteasomes was detected in the apical blebs, the apocrine secretion sites of the caput epididymal epithelia of both the rat and mouse epididymal epithelium, although region-specific differences exist. Secretion of eGFP and proteasomes continued during the prolonged culture of the isolated rat epididymal epithelial cells in vitro. This study provides evidence that the activity of the ubiquitin system is not limited to the intracellular environment, contributing to a greater understanding of the sperm maturation process during epididymal passage.
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PMID:Mechanism of extracellular ubiquitination in the mammalian epididymis. 1806 99

BACKGROUND Sperm aster organization during bovine and human fertilization requires a paternally-derived centriole that must first disengage from the sperm tail connecting-piece. We investigated the participation of the 26S proteasome in this process. METHODS Proteasome localization and enzymatic activity were studied in normal and pathological human spermatozoa by immunocytochemistry and enzyme-substrate assays. The role of proteasomes during bovine zygote development was investigated using a pharmacological proteasome-inhibitor, MG132, and with anti-proteasome antibodies delivered by Streptolysin O-permeabilization or with the Chariot reagent. Human zygotes discarded after ICSI failures (n = 28) were also examined. RESULTS Proteasomes were localized in the sperm acrosome and connecting-piece, as well as in the pronuclei of bovine and human zygotes. Proteasomal enzymatic activities were decreased in defective human spermatozoa. Disrupted sperm aster formation and pronuclear development were found after pharmacological and immunological block of proteasomes in human/bovine spermatozoa and oocytes, as well as in 28 discarded human post-ICSI fertilization failures. CONCLUSIONS Specific proteasome inhibition disrupts sperm aster formation and pronuclear development/apposition in bovine and human zygotes. Human spermatozoa with defective centriolar/pericentriolar structures have decreased proteasomal enzymatic activity. Release of a functional sperm centriole that acts as a zygote microtubule-organizing center probably relies on selective proteasomal proteolysis. These findings suggest an important role of sperm proteasomes in zygotic development.
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PMID:The role of sperm proteasomes during sperm aster formation and early zygote development: implications for fertilization failure in humans. 1808 54

Sperm-associated proteasomes have been suggested to play an important role during fertilization in animals. To delineate the role of these proteasomes during fertilization in humans, the present study reports proteasomal proteolytic activity both in noncapacitated and capacitated human spermatozoa, which is not altered in the presence of baculovirus-expressed recombinant human zona pellucida glycoprotein-3 (ZP3) and zona pellucida glycoprotein-4 (ZP4). However, inhibition of proteasomal proteolytic activity by clasto-lactacystin beta-lactone (CLBL) and Z-Leu-Leu-Leu-CHO (MG132), which are specific inhibitors of the 20S proteasomal core proteases, led to a significant (P < 0.05) inhibition of induction of acrosome reaction mediated by both recombinant human ZP3 and ZP4. Both inhibitors, however, failed to inhibit the induction of acrosomal exocytosis mediated by pharmacological agonist, calcium ionophore (A23187). The binding of recombinant human ZP3 and ZP4, labelled with fluorescein isothiocyanate, to the capacitated spermatozoa was not affected in the presence of proteasomal inhibitors. These observations suggest a role of the sperm proteasome in the induction of ZP3- and ZP4-mediated acrosomal exocytosis upstream of calcium signalling in humans.
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PMID:Role of proteasomal activity in the induction of acrosomal exocytosis in human spermatozoa. 1833 63

Proteomic profiling of the mouse spermatozoon has generated a unique and valuable inventory of candidates that can be mined for potential contraceptive targets and to further our understanding of the PTMs that regulate the functionality of this highly specialized cell. Here we report the identification of 858 proteins derived from mouse spermatozoa, 23 of which demonstrated testis only expression. The list contained many proteins that are known constituents of murine spermatozoa including Izumo, Spaca 1, 3, and 5, Spam 1, Zonadhesin, Spesp1, Smcp, Spata 6, 18, and 19, Zp3r, Zpbp 1 and 2, Spa17, Spag 6, 16, and 17, CatSper4, Acr, Cylc2, Odf1 and 2, Acrbp, and Acrv1. Certain protein families were highly represented in the proteome. For example, of the 42 gene products classified as proteases, 26 belonged to the 26S-proteasome. Of the many chaperones identified in this proteome, eight proteins with a TCP-1 domain were found, as were seven Rab guanosine triphosphatases. Finally, our list yielded three putative seven-transmembrane proteins, two of which have no known tissue distribution, an extragenomic progesterone receptor and three unique testis-specific kinases all of which may have some potential in the future regulation of male fertility.
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PMID:The mouse sperm proteome characterized via IPG strip prefractionation and LC-MS/MS identification. 1834 Jun 33


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