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Differential activity and synthesis of lactate dehydrogenase isozymes A (muscle), B (heart), and C (testis) in mouse spermatogenic cells 总被引:4,自引:0,他引:4
S S Li D A O'Brien E W Hou J Versola D L Rockett E M Eddy 《Biology of reproduction》1989,40(1):173-180
Spermatogenic cells isolated from adult and prepubertal mice by unit gravity sedimentation were used to examine enzyme activities and synthesis of the lactate dehydrogenase (LDH) isozymes during spermatogenesis. The synthesis and activity of LDH-C4, the germ cell-specific isozyme, was detected earliest in isolated preleptotene and leptotene/zygotene spermatocytes prior to the mid-pachytene stage of meiosis reported previously. The LDH-C4 isozyme was prominent in pachytene spermatocytes, round spermatids, and condensing spermatids, whereas spermatozoa contained only the LDH-C4 isozyme. In addition, somatic-type LDH isozymes consisting primarily of LDH-B subunits were present in germ cells throughout spermatogenesis. This is in contrast to a previous report that the LDH-B subunit was not synthesized in germ cells. Sertoli cells were further shown to exhibit comparable amounts of five tetrameric LDH isozymes formed by combination of muscle-type LDH-A and heart-type LDH-B subunits. 相似文献
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Drosophila TGIF is essential for developmentally regulated transcription in spermatogenesis 总被引:5,自引:0,他引:5
Ayyar S Jiang J Collu A White-Cooper H White RA 《Development (Cambridge, England)》2003,130(13):2841-2852
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Q. -L. Zhang M. Yang Y. -Y. Zhao S. -Z. Zhang Q. -H. He X. -Y. Meng W. -R. Tan 《Molecular Biology》2014,48(1):105-112
Lactate dehydrogenase C4 (LDH-C4) is considered to be a good target protein for the development of contraceptive drugs. To develop contraceptive rodenticide against pika (Ochotona curzoniae) LDH-C4, the pika LDH-C gene was cloned and expressed in Escherichia coli. The recombinant protein was purified and characterized. The cDNA of pika LDH-C gene was cloned by the RACE method. The cDNA was 1498 bp in length and contained an ORF of 996 bp, which encoded a polypeptide of 332 amino acids. The ORF of pika LDH-C was introduced in E. coli and expressed with no fusion tags added. The recombinant LDH-C4 protein was purified by heating, affinity chromatography and ion-exchange chromatography. The recombinant pika LDH-C4 was a tetramer with a molecular weight of approximately 140 kDa, and it had temperature-dependent catalytic activity, as it was thermally stable up to 60°C. The optimal pH values in the forward and backward reactions were around 7.48 and 10.28, respectively. The apparent Michaelis constants for pyruvate and lactate were 51.2 ± 3.8 and 8568.8 ± 409 μM respectively. The inhibition constant for oxalic acid was 11.8 ± 3.5 mM. This study laid a solid foundation for contraceptive rodenticide development against pika LDH-C4. 相似文献
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The consequences of error during meiotic division in spermatogenesis can be serious: aneuploid spermatozoa, embryonic lethality, and developmental abnormalities. Recombination between homologs is essential to ensure normal segregation; thus the spermatocyte must time division precisely so that it occurs after recombination between chromosomes and accumulation of the cell-cycle machinery necessary to ensure an accurate segregation of chromosomes. We use two systems to investigate meiotic division during spermatogenesis in the mouse: pharmacological induction of meiotic metaphase in cultured spermatocytes and transillumination-mediated dissection of stage XII seminiferous tubule segments to monitor progress through the division phase. By these approaches we can assess timing of acquisition of competence for the meiotic division phase and the temporal order of events as division proceeds. Competence for the meiotic division arises in the mid-pachytene stage of meiotic prophase, after chromosomes have synapsed and coincident with the accumulation of the cell-cycle regulatory protein CDC25C. The activity of both MPF and topoisomerase II are required. The earliest hallmarks of the division phase are nuclear envelope breakdown, followed by phosphorylation of histone H3 and chromosome condensation. These events are likely to be monitored by checkpoint mechanisms since checkpoint proteins can be localized in nuclei and DNA-damaging agents delay entry into the meiotic division phase. Understanding how the spermatocyte regulates its entry into the meiotic division phase can help clarify the natural mechanisms ensuring accurate chromosome segregation and preventing aneuploidy. J. Exp. Zool. (Mol. Dev. Evol.) 285:243-250, 1999. 相似文献
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A novel testis-specific protein, termed LRTP, was identified by screening both human and mouse testis and mouse pachytene spermatocyte cDNA libraries. Sequence analyses (GenBank accession number: AF092208) revealed that LRTP contains an amino terminus leucine-rich repeat domain. There are several acidic regions rich in glutamic acid in the C-terminus. The sequence, by GenBank search, shows similarities to LANP and SDS22+, leucine-rich repeat proteins localized to the nucleus and involved in the regulation of protein phosphatases. In mouse, the mRNA is first detected at about Day 14 postpartum, presumably when mid-pachytene spermatocytes are first seen. In situ hybridization confirmed the expression of the LRTP mRNA at this stage of spermatogenesis. Immunohistochemical analysis revealed that the protein is most abundant in the cytoplasm of pachytene and diplotene cells, corresponding to late prophase of meiosis I. Immunohistochemical localization is markedly reduced in secondary spermatocytes, suggesting a functional association of LRTP with meiosis. An LRTP cDNA probe did not bind to mouse ovary RNA in a dot blot assay. 相似文献
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G. S. Gupta 《Molecular and cellular biochemistry》2012,371(1-2):115-127
The lactate dehydrogenase (LDH) has been studied widely because it exists in various isozymic forms. The association of A and B subunits of LDH can generate five tetrameric isozymes, but the finding of the sixth isozyme in mature human testis and sperm indicated the presence of an additional subunit of LDH, designated as LDH-X (also termed LDH-C4 due to tetrameric nature of C-subunit). LDH-C4 isozyme is an iso-, allo-, and auto-antigen present in mammalian sperm cells. The synthesis of LDH-C4 in the testis takes place during sexual maturation, and it is the predominant fraction in mature spermatozoa. Though, originally considered to be testis specific, LDH-C or Ldh3 in mice was later detected in the murine oocyte and early embryo. Ldh3 in mouse supports its role in energy production in spermatids that favor lactate as substrate and in spermatozoa with a characteristic aerobic glycolytic path to yield ATP. During last two decades, cancer/testis-associated genes (CTAs) which are expressed only in the germinal epithelium of the testis are also expressed in some cancer cells, but not in non-cancerous somatic tissues. The CTAs are considered promising candidates for diagnosis and immunotherapy of cancer. The sperm-specific Ldh-c gene has been shown to express in a broad spectrum of human tumors, with high frequency in lung cancer, melanoma, and breast cancer; the protein being expressed virtually in all tumor types tested. Accordingly, LDH-C4 is the unique target for contraception in both males and females and offers potential future for immunotherapy of different types of cancers. As LDH-C has a preference for lactate as a substrate, LDH-C activation in cancer may depend on lactate for ATP production. The major aim of this article is to review the salient features of LDH-C subunit and the immune responses of LDH-C4 in homologous and heterologous species in relation to its role in acceptance or rejection of the allograft and its application in contraception and immunotherapy of cancer, directly or indirectly through the regulation of its substrate, the lactate. 相似文献
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Cytoplasmic events in human meiotic arrest as revealed by immunolabelling of spermatocyte proacrosin 总被引:1,自引:0,他引:1
Denise Escalier Diego Bermùdez Jean-Marc Gallo Annick Viellefond Joseph Schrével 《Differentiation; research in biological diversity》1992,51(3):233-243
Abstract. Proacrosin appears in the Golgi complex as early as the mid-pachytene stage and immediately thereafter initiates partition to be equally distributed in sper-matids. The anti-proacrosin monoclonal antibody 4D4 (mAb 4D4) was used as a marker of these cytoplasmic events in ten men exhibiting spermatogenesis arrest in three specific stages: (i) leptotene/zygotene spermatocyte I with impaired chromosome pairing (six cases), (ii) early pachytene I (one case) and (iii) metaphase/anaphase I (three cases). Prophase arrest stages were identified on testis sections stained by silver nitrate. MAb 4D4 labelling revealed two types of leptotene/zygotene arrest depending on whether proacrosin was expressed or not. The data obtained enabled us to distinguish between: (i) nuclear blockages due to chromosome and/or nuclear matrix anomalies, when cytoplasmic events were either inhibited or not inhibited, and (ii) nuclear anomalies due to microtubular disturbances. In this latter case, cytokinesis was impaired as early as the prophase I, thus indicating a relationship between the Golgi partitioning and the microtubule network. Data show that meiotic arrest investigations, by means of an appropriate marker of the cytoplasmic events, provide valuable information on spermatogenic developmental processes. 相似文献
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Spermiogenesis initiation in Caenorhabditis elegans involves a casein kinase 1 encoded by the spe-6 gene 总被引:1,自引:0,他引:1
Immature spermatids from Caenorhabditis elegans are stimulated by an external activation signal to reorganize their membranes and cytoskeleton to form crawling spermatozoa. This rapid maturation, termed spermiogenesis, occurs without any new gene expression. To better understand this signal transduction pathway, we isolated suppressors of a mutation in the spe-27 gene, which is part of the pathway. The suppressors bypass the requirement for spe-27, as well as three other genes that act in this pathway, spe-8, spe-12, and spe-29. Eighteen of the suppressor mutations are new alleles of spe-6, a previously identified gene required for an early stage of spermatogenesis. The original spe-6 mutations are loss-of-function alleles that prevent major sperm protein (MSP) assembly in the fibrous bodies of spermatocytes and arrest development in meiosis. We have isolated the spe-6 gene and find that it encodes a predicted protein-serine/threonine kinase in the casein kinase 1 family. The suppressor mutations appear to be reduction-of-function alleles. We propose a model whereby SPE-6, in addition to its early role in spermatocyte development, inhibits spermiogenesis until the activation signal is received. The activation signal is transduced through SPE-8, SPE-12, SPE-27, and SPE-29 to relieve SPE-6 repression, thus triggering the formation of crawling spermatozoa. 相似文献
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Conservation of the heterochronic regulator Lin-28, its developmental expression and microRNA complementary sites 总被引:14,自引:0,他引:14
The heterochronic gene lin-28 is a regulator of developmental timing in the nematode Caenorhabditis elegans. It must be expressed in the first larval stage and downregulated by the second stage for normal development. This downregulation is mediated in part by lin-4, a 21-nt microRNA. If downregulation fails due to a mutation in a short sequence in the lin-28 3' UTR that is complementary to lin-4, then a variety of somatic cell lineages fail to progress normally in development. Here, we report that Lin-28 homologues exist in diverse animals, including Drosophila, Xenopus, mouse, and human. These homologues are characterized by the LIN-28 protein's unusual pairing of RNA-binding motifs: a cold shock domain (CSD) and a pair of retroviral-type CCHC zinc knuckles. Conservation of LIN-28 proteins shows them to be distinct from the other conserved family of CSD-containing proteins of animals, the Y-box proteins. Importantly, the LIN-28 proteins of Drosophila, Xenopus, and mouse each appear to be expressed and downregulated during development, consistent with a conserved role for this regulator of developmental timing. In addition, the extremely long 3' UTRs of mouse and human Lin-28 genes show extensive regions of sequence identity that contain sites complementary to the mammalian homologues of C. elegans lin-4 and let-7 microRNAs, suggesting that microRNA regulation is a conserved feature of the Lin-28 gene in diverse animals. 相似文献
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Lee B Park I Jin S Choi H Kwon JT Kim J Jeong J Cho BN Eddy EM Cho C 《The Journal of biological chemistry》2011,286(33):29108-29117
Spermatogenesis is a complex process involving an intrinsic genetic program composed of germ cell-specific and -predominant genes. In this study, we investigated the mouse Spink2 (serine protease inhibitor Kazal-type 2) gene, which belongs to the SPINK family of proteins characterized by the presence of a Kazal-type serine protease inhibitor-pancreatic secretory trypsin inhibitor domain. We showed that recombinant mouse SPINK2 has trypsin-inhibitory activity. Distribution analyses revealed that Spink2 is transcribed strongly in the testis and weakly in the epididymis, but is not detected in other mouse tissues. Expression of Spink2 is specific to germ cells in the testis and is first evident at the pachytene spermatocyte stage. Immunoblot analyses demonstrated that SPINK2 protein is present in male germ cells at all developmental stages, including in testicular spermatogenic cells, testicular sperm, and mature sperm. To elucidate the functional role of SPINK2 in vivo, we generated mutant mice with diminished levels of SPINK2 using a gene trap mutagenesis approach. Mutant male mice exhibit significantly impaired fertility; further phenotypic analyses revealed that testicular integrity is disrupted, resulting in a reduction in sperm number. Moreover, we found that testes from mutant mice exhibit abnormal spermatogenesis and germ cell apoptosis accompanied by elevated serine protease activity. Our studies thus provide the first demonstration that SPINK2 is required for maintaining normal spermatogenesis and potentially regulates serine protease-mediated apoptosis in male germ cells. 相似文献
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Stage-dependent appearance of sulfhydryl oxidase during spermatogenesis in the testis of rat and hamster. An immunohistochemical study 总被引:1,自引:0,他引:1
Sulfhydryl oxidase (SOx), an enzyme that catalyzes the oxidation of sulfhydryl compounds, appears in the spermatogenic cells of rat and hamster testes in a stage-dependent manner. It first appears in pachytene spermatocytes at stage I in both the animal species studied. SOx immunoreactivity is associated with mitochondria of these cells. The fate of such mitochondria is species-dependent. In rat, the immunoreactive mitochondria aggregate during maturation phase and are retained in the residual bodies. Spermatozoa free of SOx are released into the lumen. On the other hand, in hamster, the immunoreactive mitochondria arrange themselves around the midpiece of spermatozoa. In such a case, residual bodies lack SOx. The appearance of SOx coincides with the appearance of LDH-X in the spermatogenic cells. Like many other proteins such as LDH-X, RSA-1 and cytochrome ct, SOx provides yet another example of differential gene activation associated with a developmental process of gametes. 相似文献