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)

Ubiquitination of the sperm mitochondria during spermatogenesis has been implicated in the targeted degradation of paternal mitochondria after fertilization, a mechanism proposed to promote the predominantly maternal inheritance of mitochondrial DNA in humans and animals. The identity of ubiquitinated substrates in the sperm mitochondria is not known. In the present study, we show that prohibitin, a highly conserved, 30- to 32-kDa mitochondrial membrane protein, occurs in a number of unexpected isoforms, ranging from 64 to greater than 185 kDa in the mammalian sperm mitochondria, which are the ubiquitinated substrates. These bands bind antiubiquitin antibodies, displaying a pattern consistent with polyubiquitinated "ladders." Immunoprecipitation of sperm extracts with antiprohibitin antibodies followed by probing of the resultant immunocomplexes with antiubiquitin yields a banding pattern identical to that observed by antiprohibitin Western blot analysis. In fact, the presumably nonubiquitinated 30-kDa prohibitin band shows no antiubiquitin immunoreactivity. We demonstrate that ubiquitination of prohibitin occurs in testicular spermatids and spermatozoa. Ubiquitinated prohibitin molecules also accumulate in the defective fractions of ejaculated spermatozoa, which are thought to undergo surface ubiquitination during epididymal passage. In such sperm fractions, ubiquitin also coprecipitates with tubulin and microtubule-associated proteins, presumably contributed by the axonemes of defective, ubiquitinated spermatozoa. The results of the present study suggest that prohibitin is one of the ubiquitinated substrates that makes the sperm mitochondria recognizable by the egg's ubiquitin-proteasome dependent proteolytic machinery after fertilization and most likely facilitates the marking of defective spermatozoa in the epididymis for degradation.
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PMID:Ubiquitination of prohibitin in mammalian sperm mitochondria: possible roles in the regulation of mitochondrial inheritance and sperm quality control. 1264 88

Ubiquitin and ubiquitin-like proteins control the degradation of substrates as diverse as cyclins, viral envelope proteins, plasma membrane receptors, and mRNAs. The ubiquitinated substrates are targeted towards the lysosomal or proteasomal degradation sites. The number and position of ubiquitin molecules bound to substrates' lysine residues and the number and position of ubiquitin molecules in polyubiquitin chains determine the astonishing substrate specificity of ubiquitin-mediated proteolysis. Ubiquitin is likely to be expressed in mammalian gametes and embryos at any given developmental step, but the information on ubiquitin dependence of gametogenesis and fertilization is sketchy. Ubiquitin ligases E1, E2, E3, and UBC4 are active in the testis. Ubiquitin and proteasomal subunits can be detected in the human sperm centrosome that undergoes dramatic reduction during spermatid elongation. Spermatid histones are ubiquitinated as they are being transiently replaced by transitional proteins and permanently by protamines. Ubiquitin tagging of the sperm mitochondrial membranes may serve as a death sentence for paternal mitochondria at fertilization, thus promoting the maternal inheritance of mitochondrial DNA (mtDNA) in mammals. The defective spermatozoa become surface-ubiquitinated during sperm descent down the epididymis. Finally, new evidence suggests the involvement of ubiquitin-proteasome pathway in the zona penetration by the acrosome-reacted spermatozoon. Such differential patterns of ubiquitination in the testis and epididymis, and inside the egg, may be necessary for reproductive success in humans and animals. Deciphering and eventually manipulating the ubiquitin-dependent proteolysis in the reproductive system could allow us to redirect the mode of mtDNA inheritance after cloning and ooplasmic transplantation, provide germ line therapy in some cases of male infertility, develop new contraceptives, manage polyspermia during in vitro fertilization, and establish objective markers for infertility diagnostics, semen evaluation, and prediction of future fertility.
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PMID:Ubiquitin-dependent proteolysis in mammalian spermatogenesis, fertilization, and sperm quality control: killing three birds with one stone. 1267 25

A gene coding a novel human RAD23B protein named RAD23-like protein B, presumably involved in spermatogenesis, was identified and characterized using a complementary DNA (cDNA) microarray. In the human testis, its expression was 2.33 times higher than it was in the embryo testis, with a particularly high expression in ejaculated human spermatozoa. The full length of this gene is 1548 bp, and the putative protein is 338 amino acids long. This protein is homologous to RAD23B, which is one of two human homologs of Saccharomyces cerevisiae RAD23, and is involved in both nucleotide excision repair (NER) and ubiquitin (Ub)-dependent proteolysis. However, RAD23-like protein B lacks the Ub-like (UbL) domain that functions as a proteasome localization signal. Multiple-tissue expression profile of the messenger RNA (mRNA) that encodes the RAD23-like protein B also showed that it is highly expressed in the human testis and in ejaculated spermatozoa. Our present study indicates that this novel alternative splicing form of RAD23B is correlated with human spermatogenesis.
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PMID:Expression of a novel RAD23B mRNA splice variant in the human testis. 1506 13

A long-standing problem in epididymal physiology is the fate of unejaculated spermatozoa in the cauda epididymidis under conditions such as congenital absence of the vas deferens, long-term vasectomy, or castration. There is no convincing evidence for significant absorption of spermatozoa, defective or otherwise, by spermiophagy or dissolution in the epididymis of normal animals. Spermiophagy by epithelial cells or intraluminal macrophages may take place if the duct ruptures and granulomas form (e.g., after experimental ligation), although there is no quantitative information on the rate of sperm removal by this means. In one animal model (the rabbit), the epididymis is unusually resistant to granuloma formation and has provided unique insights into a phenomenon that is suggested to be present in all species. Spermatozoa retained in the rabbit cauda epididymidis by placing ligatures on the vas deferens and corpus epididymidis degenerate after several weeks but do not decrease significantly in numbers. After castration, however, they die very rapidly and >90% disappear. It is hypothesized that, in the normal androgen-maintained epididymis, degradative pathways are present in the luminal fluid that are constitutively inhibited by survival signals emanating from the epithelium. In the absence of androgen, the intraluminal mileau changes and death signals predominate that activate degradative pathways via the ubiquitin-proteasome system, DNAses, etc., to mediate dissolution of sperm organelles and nucleoprotein. It is suggested that the latter condition is the default situation and is only prevented by the stimulatory action of androgens on the epididymal epithelium.
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PMID:Sperm survival versus degradation in the Mammalian epididymis: a hypothesis. 1521 93

The rat analogue of the complement regulator membrane cofactor protein (MCP; CD46) was recently cloned and analysis at the mRNA level suggested that expression was restricted to testis. In light of the proposed roles of human MCP in sperm-egg interaction, we undertook to analyze rat MCP expression at the protein level in order better to address its putative role in fertilization. Recombinant fusion proteins comprising antibody Fc and specific domains of rat MCP were generated and used to develop a monoclonal antibody, MM.1, specific for rat MCP. Immunohistochemistry using these reagents confirmed the reported testis-specific expression of MCP in sexually mature rats and demonstrated that MCP was expressed only by spermatozoa and their immediate precursors in spermiogenesis, spermatids. Prepubertal male rats did not express MCP, and there was no evidence of MCP expression at any site in the embryo. Spermatozoal MCP expression was restricted to the inner acrosomal membrane, exposed only after fixation or induction of the acrosome reaction. Acrosome-reacted but not unreacted spermatozoa bound methylamine-activated C3 immobilized on plastic. The retention of MCP at this subcellular site, which is probably crucial to sperm-egg interaction, and the functional demonstration of binding to activated C3 strengthen suggestions from human studies that MCP may play an important role in fertilization. The reagents and results described here will enable studies of the role of spermatozoal MCP in sperm-egg interaction using a relevant animal model system.
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PMID:Rat membrane cofactor protein (MCP; CD46) is expressed only in the acrosome of developing and mature spermatozoa and mediates binding to immobilized activated C3. 1521 99

The mouse USP8/mUBPy gene codifies a deubiquitinating enzyme expressed preferentially in testis and brain. While the ubiquitin-specific processing proteases (UBPs) are known to be important for the early development in invertebrate organisms, their specific functions remain still unclear in mammals. Using specific antibodies, raised against a recombinant mUBPy protein, we studied mUBPy in mouse testis. The mUBPy is expressed exclusively by the germ cell component and is maintained in epididymal spermatozoa. The enzyme is functionally active, being able to detach ubiquitin moieties from endogenous protein substrates. Protein interaction assays showed that sperm UBPy interacts with MSJ-1, the sperm-specific DnaJ protein evolutionarily conserved for spermiogenesis. Immunocytochemistry revealed that mUBPy shares with MSJ-1 the intracellular localization during spermatid cell differentiation; intriguingly, we show here that the proteasomes also locate in mUBPy/MSJ-1-positive sites, such as the cytoplasmic surface of the developing acrosome and the centrosomal region. These colocalization sites are maintained in epididymal spermatozoa. The demonstration of a protein interaction between a deubiquitinating enzyme and a molecular chaperone and the documentation on the proteasomes in both differentiating and mature mouse male germ cells suggest that members of the chaperone and ubiquitin/proteasome systems could cooperate in the fine control of protein quality to yield functional spermatozoa.
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PMID:The deubiquitinating enzyme mUBPy interacts with the sperm-specific molecular chaperone MSJ-1: the relation with the proteasome, acrosome, and centrosome in mouse male germ cells. 1534 53

Sperm proteolytic activities are relevant in the enzymatic mechanism of fertilization. Several authors have suggested the presence of serine proteases other than acrosin in mice and human spermatozoa. In this work we describe the characterization of a partially purified bovine sperm serine protease BSp66 and its dimmer, BSp120. Partial purification of the monomer was performed from fresh spermatozoa, while the dimer form of the protease was obtained from cryopreserved spermatozoa. The Mr of BSp120 and BSp66 estimated by zymography and gel filtration chromatography were 120 and 66 kDa, respectively. They were positively stained by Schiff-PAS reagent for glycoproteins and they both digested synthetic peptides with basic amino acids in the P1 site. Polyclonal antibodies against acrosin or proacrosin did not cross-react neither with BSp120, nor BSp66. In addition, antibodies raised in our laboratory against BSp120 and BSp66 did not recognize acrosin or proacrosin suggesting that they are not antigenically related proteins. Also, no cross- reactivity was detected with proteins in the range of 120-66 kDa when antibodies against the proteasome were used. The cellular localization of this protease by optical immunocytochemistry using specific antibodies revealed a positive signal in the apical portion of the sperm head suggesting acrosomal or membrane localization. The evidences presented here characterize BSp66 as a trypsin-like serine protease, a putative new member of this highly redundant proteolytic system of the sperm acrosome.
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PMID:Partial purification and characterization of a trypsin-like serine protease from bovine sperm. 1537 73

The contraceptive efficacy and toxicological screening of the two principal compounds, MCP I and ECP I, isolated from the seeds of Carica papaya, in male albino rats at the standardized dose regimen, at 50 mg/kg b.w./day, for a period of 360 days and up to 90 days of treatment withdrawal have been reported. The body and organ weights, cauda epididymal sperm characteristics, androgen sensitive tissue biochemistry, reactive oxygen species and anti-oxidant defense system in the cauda epididymal microenvironment, histology and ultrastructure of testis and cauda epididymis, histology of seminal vesicle and prostate, toxicological investigations through routine hematology and serum clinical chemistry, sexual behaviour and fertility index have been studied. The results revealed that oral administration of MCP I and ECP I were equally effective, exhibiting complete inhibition of sperm motility following 90 days of treatment that coincided with a gradual and significant decline in cauda epididymal sperm density, percent viable spermatozoa and significant increase in sperm anomalies. Histology of testis of treated animals revealed degenerated germinal epithelium, vacuolization in Sertoli cells and proliferating germ cells and disturbances in spermatid differentiation. Spermatogonial stem cell reserves and Leydig cells appeared normal. Ultrastructure of the testis revealed vacuolization in the Sertoli cells and germ cells, loss of cytoplasmic characteristics in the Sertoli cells, nuclear degeneration and mitochondrial vacuolization in spermatocytes and spermatids. Leydig cells exhibited steroidogenic features. Cauda epididymis showed normal epithelial cell function. Absence of spermatozoa or disruption of spermatozoa clusters in the lumen were evident. Ultrastructure of cauda epididymis revealed normal secretory activity. Morphology of seminal vesicle and prostate of the treated animals were comparable to control animals. Serum testosterone, tissue biochemical and toxicological parameters remained unaffected. Fertility test revealed 100% efficacy. All the altered parameters showed sign of recovery following 90 days of treatment withdrawal. It is concluded that both MCP I and ECP I are equally effective in terms of contraceptive efficacy which is likely reversible and without adverse side effects.
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PMID:Efficacy trial on the purified compounds of the seeds of Carica papaya for male contraception in albino rat. 1580 97

A specific hypoglycosylated isoform of the complement regulator membrane cofactor protein (MCP; CD46) is expressed on the inner acrosomal membrane (IAM) of spermatozoa. This membrane is exposed after the acrosome reaction, an exocytosis event that occurs upon contact with the zona pellucida. We initiated this investigation to assess MCP's regulatory function in situ on spermatozoa. Upon exposure of human spermatozoa to autologous serum or follicular fluid, we unexpectedly observed that acrosome-reacted spermatozoa activated the complement cascade efficiently through C3 but not beyond. Using FACS to simultaneously evaluate viability, acrosomal status, and complement deposition, we found that complement activation was initiated by C-reactive protein (CRP) and was C1q, C2, and factor B dependent. This pattern is consistent with engagement of the classical pathway followed by amplification through the alternative pathway. C3b deposition was targeted to the IAM, where it was cleaved to C3bi. Factor H, and not MCP, was the cofactor responsible for C3b cleavage. We propose that this localized deposition of complement fragments aids in the fusion process between the spermatozoa and egg, in a role akin to that of complement in immune adherence. In addition, we speculate that this "targeted and restricted" form of complement activation on host cells is a common strategy to handle modified self.
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PMID:Targeted and restricted complement activation on acrosome-reacted spermatozoa. 1584 10

Lipoxygenases (LOXs) are a family of enzymes capable of peroxidizing phospholipids. A member of the LOX family of enzymes, 15-LOX, participates in the degradation of mitochondria and other organelles within differentiating red blood cells, the reticulocytes. The present study provides biochemical and immunocytochemical evidence for the presence of 15-LOX in the sperm cytoplasmic droplet (CD). Testicular, epididymal and ejaculated spermatozoa were evaluated for the presence of 15-LOX using an affinity-purified immune serum raised against a synthetic peptide corresponding to the C-terminal sequence of rabbit reticulocyte 15-LOX. Western blotting revealed an appropriate single band of approximately 81 kDa in boar spermatozoa but not in boar seminal plasma. When ejaculated boar spermatozoa were subjected to separation on a 45/90% Percoll gradient, 15-LOX co-migrated with the immotile sperm and cellular debris/CD fractions, but not with the motile sperm fraction containing morphologically normal spermatozoa without CDs. Varied levels of 15-LOX were expressed in ejaculated sperm samples from boars with varied semen quality. By immunofluorescence, prominent 15-LOX immunoreactivity was found within the residual body in the testis and within the CDs from caput, corpus and cauda epididymal and ejaculated spermatozoa. Components of the ubiquitin-dependent proteolytic pathway, which is thought to facilitate both spermiogenesis and reticulocyte organelle degradation, were also detected in the sperm CD. These included ubiquitin, the ubiquitin-conjugating enzyme E2, the ubiquitin C-terminal hydrolase PGP 9.5, and various 20S proteasomal core subunits of the alpha- and beta-type. The 15-LOX and various components of the ubiquitin-proteasome pathway were also detected in sperm CDs of other mammalian species, including the human, mouse, stallion and wild babirusa boar. We conclude that 15-LOX is prominently present in the mammalian sperm CD and thus may contribute to spermiogenesis, CD function or CD removal.
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PMID:15-Lipoxygenase is a component of the mammalian sperm cytoplasmic droplet. 1604 59


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