UNIPROT:P01034 - FACTA Search

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Query: UNIPROT:P01034 (cystatin C)
3,397 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Procathepsin L, the precursor to a powerful lysosomal cysteine proteinase, has been purified to apparent homogeneity from guinea pig spermatozoa, a novel and previously unrecognized source of this catalytically active zymogen. In the range of pH 5.0, procathepsin L (39,000 M(r)) readily self-processed yielding a mature, single-chain proteinase (29,000 M(r)) and an intact propeptide (10,000 M(r)) by what appeared kinetically to be an intramolecular reaction mechanism. These characteristics resembled those reported for the "major excreted protein" (MEP) of malignantly transformed mouse fibroblasts-a protein that has been characterized as the precursor to the mouse analog of human cathepsin L (B. R. Troen, S. Gal, and M. M. Gottesman (1987) Biochem. J. 246, 731-735). Other characteristics shared by the guinea pig and mouse zymogens included proteolytic activity at pH 5.0, homologous N-terminal amino acid sequences, and immunological relatedness. It was thus concluded that acrosomal procathepsin L is the guinea pig analog of MEP. Acrosomal procathepsin L had a specific activity on benzyloxy-carbonyl-Phe-Arg-7-(4-methyl)coumarylamide (Z-Phe-Arg-NMec) of 30 mumol min-1 mg-1 enzyme at pH 3.2 and 37 degrees C. Relative to the assay substrate, rates on other fluorogenic substrates were 90% for Z-Phe-Cit-NMec, 63% for Z-Leu-Leu-Arg-NMec, 43% for D-Phe-Ser(Bzl)-Phe-Phe-Ala-Ala-p-aminobenzoate (a "specific" cathepsin D assay substrate), and 32% for Z-Val-Val-Arg-NMec. No action was detected on Z-Arg-Arg-NMec or Arg-NMec. Mature cathepsin L showed the same relative order of substrate specificity as its proenzyme form, but the absolute rates were about 5-fold greater. Additionally, the mature (single-chain) form of cathepsin L displayed Km and kcat values on Z-Phe-Arg-NMec that yielded an exceptionally high catalytic coefficient (11,600 s-1 mM-1) compared to values reported for two-chain forms of cathepsin L. Self-processing by acrosomal procathepsin L at pH 5.5 was totally inhibited by leupeptin, cystatin C, Ep-475, and Z-Phe-Phe-CHN2 at 1 microM levels. Gossypol (0.1 mM) gave 94% inhibition. Interestingly, dextran sulfate (100 micrograms ml-1) gave a 3.6-fold increase in the rate of self-processing seen at pH 5.5--a phenomenon of potential physiological relevance in view of the high-negative-charge density present within the hyaluronic acid-rich outer layer (cumulus oophorus) of the ovum.
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PMID:Purification and characterization of procathepsin L, a self-processing zymogen of guinea pig spermatozoa that acts on a cathepsin D assay substrate. 748 6

The specific inhibitor of cysteine proteinases, cystatin C, was purified from ram rete testis fluid and the conditioned medium of Sertoli cells. This molecule associated with sheep liver cathepsin L at one of the fastest rates ever described for a proteinase/inhibitor interaction (1.75 +/- 0.20 x 10(8) M-1.s-1). But the association rate constant for the interaction of cathepsin L with alpha 2-macroglobulin, a non-specific inhibitor of proteinases, was also extremely high (8.8 +/- 0.75 x 10(6) M-1.s-1). Cathepsin L complexed with alpha 2-macroglobulin was protected from inhibition by type 2 and type 3 cystatins. The data indicate that cystatin C is the most potent inhibitor of cathepsin L in mammalian male genital tract fluids, whereas alpha 2-macroglobulin may act as a terminal acceptor of this enzyme. These inhibitors could therefore inhibit the activated form of procathepsin L which may appear during the complex process of spermatozoa production and maturation in the testis and epididymis.
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PMID:Interactions between ovine cathepsin L, cystatin C and alpha 2-macroglobulin. Potential role in the genital tract. 906 57

Cystatins are cysteine proteinase inhibitors. We found two expression sequence tags (ESTs), CA463109 and AV042522, from a mouse testis library using Digital differential display (DDD). By electrical hybridization, a novel gene, Cymg1 (GenBank accession No. AY600990), which has a full length of 0.78 kb, and contains four exons and three introns, was cloned from a mouse testis cDNA library. The gene is located in the 2G3 area of chromosome 2. The full cDNA encompasses the entire open reading frame, encoding 141 amino acid residues. The protein has a cysteine protease inhibitor domain that is related to the family 2 cystatins but lacks critical consensus sites important for cysteine protease inhibition. These characteristics are seen in the CRES subfamily, which are related to the family 2 cystatins and are expressed specifically in the male reproductive tract. CYMG1 has a 44% (48/108) identity with mouse CRES and 30% (42/140) identity with mouse cystatin C. Northern blot analysis showed that the Cymg1 is specifically expressed in adult mouse testes. Cell location studies showed that the GFP-tagged CYMG1 protein was localized in the cytoplasm of HeLa cells. Immunohistochemistry revealed that the CYMG1 protein was expressed in mouse testes spermatogonium, spermatocytes, round spermatids, elongating spermatids and spermatozoa. RT-PCR results also showed that Cymg1 was expressed in mouse testes and spermatogonium. The Cymg1 expression level varied in different developmental stages: it was low 1 week postpartum, steadily increased 2 to 5 weeks postpartum, and was highest 7 weeks postpartum. The expression level at 5 weeks postpartum was maintained during 13 to 57 weeks postpartum. The Cymg1 expression level in the testes over different developmental stages correlates with the mouse spermatogenesis and sexual maturation process. All these indicate that Cymg1 might play an important role in mouse spermatogenesis and sexual maturation.
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PMID:Cloning, characterization and primary function study of a novel gene, Cymg1, related to family 2 cystatins. 1564 76

In this study, a homology model of carp ovum cystatin was constructed based on the crystal structure of chicken egg white cystatin. The results of amino acid sequence alignment indicate that these two proteins exhibit 36.11% of sequence identity. The resultant homology model reveals that carp ovum cystatin shares similar folds as chicken egg white cystatin, particularly in the conserved regions of Q48-V49-G52 and P98-W99 and the locations of two disulfide bonds, C67-C76 and C90-C110. However, the results of 1 ns molecular dynamics simulations show that carp ovum cystatin exhibits less structural integrity than chicken egg white cystatin in explicit water at 300 K. The relatively hydrophilic Met62 of carp ovum cystatin, corresponding to the hydrophobic Leu68 of human cystatin C and Ile66 of chicken egg white cystatin, may destabilize the hydrophobic core and form a dimeric structure more easily through domain swapping. A total of 16 positively charged residues are equally distributed on the surface of carp ovum cystatin, resulting in agglutination with the negatively charged spermatozoa via electrostatic interaction. Thus, carp ovum cystatin is considered to be important in preventing carp eggs from polyspermy.
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PMID:Homology model and molecular dynamics simulation of carp ovum cystatin. 1608 Jul 17

The epididymal lumen represents a unique extracellular environment because of the active sperm maturation process that takes place within its confines. Although much focus has been placed on the interaction of epididymal secretory proteins with spermatozoa in the lumen, very little is known regarding how the complex epididymal milieu as a whole is maintained, including mechanisms to prevent or control proteins that may not stay in their native folded state following secretion. Because some misfolded proteins can form cytotoxic aggregate structures known as amyloid, it is likely that control/surveillance mechanisms exist within the epididymis to protect against this process and allow sperm maturation to occur. To study protein aggregation and to identify extracellular quality control mechanisms in the epididymis, we used the cystatin family of cysteine protease inhibitors, including cystatin-related epididymal spermatogenic and cystatin C as molecular models because both proteins have inherent properties to aggregate and form amyloid. In this chapter, we present a brief summary of protein aggregation by the amyloid pathway based on what is known from other organ systems and describe quality control mechanisms that exist intracellularly to control protein misfolding and aggregation. We then present a summary of our studies of cystatin-related epididymal spermatogenic (CRES) oligomerization within the epididymal lumen, including studies suggesting that transglutaminase cross-linking may be one mechanism of extracellular quality control within the epididymis.
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PMID:Extracellular quality control in the epididymis. 1758 87

CRES (cystatin-related epididymal spermatogenic), a member of the cystatin superfamily of cysteine protease inhibitors, is expressed in the epididymis and spermatozoa, suggesting specialized roles in reproduction. Several cystatin family members oligomerize, including cystatin C that forms amyloid deposits associated with cerebral amyloid angiopathy. Our studies demonstrate that CRES also forms oligomers. Size exclusion chromatography revealed the presence of multiple forms of CRES in the epididymal luminal fluid, including SDS-sensitive and SDS-resistant high molecular mass complexes. In vitro experiments demonstrated that CRES is a substrate for transglutaminase and that an endogenous transglutaminase activity in the epididymal lumen catalyzed the formation of SDS-resistant CRES complexes. The use of a conformation-dependent antibody that recognizes only the oligomeric precursors to amyloid, negative stain electron microscopy, and Congo Red staining showed that CRES adopted similar oligomeric and fibrillar structures during its aggregation as other amyloidogenic proteins, suggesting that CRES has the potential to form amyloid in the epididymal lumen. The addition of transglutaminase, however, prevented the formation of CRES oligomers recognized by the conformation antibody by cross-linking CRES into an amorphous structure. We propose that transglutaminase activity in the epididymal lumen may function as a mechanism of extracellular quality control by diverting proteins such as CRES from the amyloidogenic pathway.
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PMID:Oligomerization and transglutaminase cross-linking of the cystatin CRES in the mouse epididymal lumen: potential mechanism of extracellular quality control. 1785 42

Polyploidy, hybridization and variation in mating systems are central issues for a deeper understanding of animal evolution. The Iberian species Squalius alburnoides represents an example combining all three phenomena. Previous studies showed that S. alburnoides populations are mainly composed of triploid and diploid hybrid forms (mainly females), and that the tetraploid forms are rare or absent. Both populations from the Douro drainage reveal a distinct scenario: tetraploid individuals represent 85.6-97.5% of the population, with no sex ratio bias observed. Based on the flow cytometry measurements of blood and spermatozoa cells, microsatellite loci and experimental crosses, we describe here, for the first time, two symmetric allotetraploid populations (CCAA) that resumed normal meiosis after undergoing intermediate processes of non-sexual reproduction to give rise to a new sexually reproducing polyploid species. Prezygotic (habitat selection and assortative mating) and postzygotic mechanisms (nonviable embryos) are responsible for the reproductive isolation from other forms of the S. alburnoides complex (e.g. CA, CAA). This example illustrates how hybrid polyploid complexes may lead to speciation.
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PMID:Speciation towards tetraploidization after intermediate processes of non-sexual reproduction. 1852 19

The spinocerebellar ataxias (SCA) are a genetically and clinically heterogeneous group of diseases, characterized by dominant inheritance, progressive cerebellar ataxia and diverse extracerebellar symptoms. A subgroup of the ataxias is caused by unstable CAG-repeat expansions in their respective genes leading to pathogenic expansions of polyglutamine stretches in the encoded proteins. In general, unstable CAG repeats have an uninterrupted CAG repeat, whereas stable CAG repeats are either short or interrupted by CAA codons, which - like CAG codons - code for glutamine. Here we report on an infantile SCA2 patient who, due to germ-line CAG repeat instability in her father, inherited an extremely expanded CAG repeat in the SCA2 locus. Surprisingly, the expanded allele of the father was an interrupted CAG repeat sequence. Furthermore, analyses of single spermatozoa showed a high frequency of paternal germ-line repeat sequence instability of the expanded SCA2 locus.
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PMID:Germ-line CAG repeat instability causes extreme CAG repeat expansion with infantile-onset spinocerebellar ataxia type 2. 2304 44

Hereditary cystatin C amyloid angiopathy is an autosomal dominant disorder in which a variant form of cystatin C (L68Q) readily forms amyloid deposits in cerebral arteries in affected individuals resulting in early death. L68Q protein deposits in human cystatin C amyloid angiopathy patients have also been found in tissues outside of the brain including the testis, suggesting possible effects on fertility. Heterozygous transgenic mice (L68Q) that express the human L68Q variant of cystatin C under the control of the mouse cystatin C promoter were unable to generate offspring, suggesting the presence of L68Q cystatin C amyloid affected sperm function. In vitro studies showed that epididymal spermatozoa from L68Q mice were unable to fertilize oocytes and exhibited poor sperm motility. Furthermore, spermatozoa from L68Q mice exhibited reduced cell viability compared with wild type (WT) spermatozoa and often were detected in large agglutinated clumps. Examination of the epididymal fluid and spermatozoa from L68Q mice showed increased levels and distinct forms of cystatin C amyloid that were not present in WT mice. The addition of epididymal fluid from L68Q mice to WT spermatozoa resulted in a recapitulation of the L68Q phenotype in that WT spermatozoa showed reduced cell viability and motility compared with WT spermatozoa incubated in epididymal fluid from WT mice. L68Q epididymal fluid that was depleted of cystatin C amyloids, however, did not impair the motility of WT spermatozoa. Taken together these studies suggest that amyloids in the epididymal fluid can be cytotoxic to the maturing spermatozoa resulting in male infertility.
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PMID:Fertility defects in mice expressing the L68Q variant of human cystatin C: a role for amyloid in male infertility. 2450 Jul 19

Immature spermatozoa undergo series of events in the epididymis to acquire motility and fertilizing ability. These events are a direct result of exposure to, and interaction with, the luminal environment created by the epididymal epithelium. The three conventional regions of the epididymis namely; caput, corpus and cauda have been identified to play specific roles in the epididymal maturation process of the spermatozoa; their respective roles have been associated with specific gene expression patterns that account for the composition of the luminal fluid that bathe the spermatozoa as they transit through the epididymal lumen and ensure their maturation. The identification of genes expressed in a region-specific manner provides valuable insight into the functional differences among the regions. Microarray technology has previously been employed in region-specific gene expression studies using the epididymis as a model in different species such as mouse, rat, boar and human. However, to characterize gene expression in the different regions of the epididymis, RNA-seq analysis was used in our study to examine gene expressions in the caput, corpus, and cauda of yak epididymis. Comparative transcriptomic analysis was performed between region pairs in the order; caput vs corpus, caput vs cauda and corpus vs cauda. DEGs among the various region pairs were detected and functional analysis were performed for the detected DEGs. Overall, the caput vs cauda epididymidis pair produced the highest number of DEGs (49.4%) while the corpus vs cauda pair produced the least number of DEGs (19.3%). The caput segment demonstrated relatively high expression of Sal1, LCN6, PTDS, DEFB109, DEFB 119, DEFB 123, SPAG11, PROC, CST3, ADAM28, KCNJ12 and SLC13A2; corpus epididymis demonstrated relatively high expression of MAN2B2, ELP, ZFYVE21, GLB1L, BMP4, DEFB125, PPP1R10, RIOX2, TKDP1, DEFB106A, NPBWR1 and SLC28A1; and the cauda epididymis, demonstrated relatively high expressions of MCT7, PAG4, OAS1, TGM3 and PRSS45. Gene Ontology results showed that DEGs in the caput vs corpus and corpus vs cauda pairs were mostly enriched in the cell/cell part GO term. On the other hand, DEGs in the caput vs cauda pair was were mostly enriched in the cellular process term. KEGG pathway annotation was also performed for DEGs among the various groups. AMPK signaling pathway, which is characterized by the ratio between cellular AMP and ATP and also determines cellular energy state, was selected from among the top five KEGG pathways for DEGs in the caput vs corpus pair. Our results showed that some down-regulated DEGs in the caput and corpus pair such as HN4a, eEF2K and CFTR were present and played significant roles in the AMPK signaling pathway. In the corpus vs cauda pair, our results showed that up-regulated DEGs such as XDH, TRMP2 and ENTPD were involved in the purine metabolism KEGG pathway, which was among top five KEGG pathways for DEGs in this pair. Pentose phosphate pathway functions in antioxidation to protect both the spermatozoa and epididymis from oxidative damage; it was among top five KEGG pathways for DEGs in the caput vs cauda pair. Our results also showed that down-regulated genes in the caput vs cauda pair such as TALDO1 was found to be involved in the Pentose phosphate pathway. The significance of the upregulated and downregulated genes on the pathways were elucidated. SAL1, which showed high expression in the caput, had previously not been demonstrated in the epididymis, needs further investigation to establish its unique role in the yak epididymis.
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PMID:Region-specific gene expression in the epididymis of Yak. 3140 23

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