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Margarita Vigodner
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ABSTRACT: Spermatogenesis consists of the mitotic division of spermatogonia, meiosis of spermatocytes, and postmeiotic differentiation of spermatids, processes tightly controlled by hormones and growth factors secreted by testicular somatic cells. The events during spermatogenesis are precisely regulated by the sequential appearance of different proteins and their posttranslational modifications. Sumoylation (covalent modification by small ubiquitin-like modifiers; SUMO proteins) has emerged as an important regulatory mechanism in different cell types, and data obtained from studies on germ cells imply that SUMO proteins are involved in multiple aspects of spermatogenesis. Although progress has been made in the initial characterization of sumoylated proteins during spermatogenesis, the targets of sumoylation, their corresponding pathways in the testis, are mostly unknown. In this chapter, I review what we know about sumoylation in somatic cells, summarize the expression patterns, suggest possible functions of SUMO proteins in testicular cells, and discuss some difficulties and perspectives on the studies of sumoylation during spermatogenesis.
International review of cell and molecular biology 01/2011; 288:227-59. · 4.48 Impact Factor
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ABSTRACT: Small ubiquitin-like modifiers (SUMO) proteins have been implicated in cellular stress response in different tissues, but whether sumoylation has a similar role during spermatogenesis is currently unknown. In this study, changes in the levels of both free SUMO isoforms and high-molecular weight (HMW) SUMO conjugates were monitored before and after the induction of different types of cellular stresses. Using cell lines and primary cells freshly isolated from mouse testes, significant changes were detected in the levels of SUMO1 and SUMO2/3 conjugates following short exposure of the cells to heat stress and oxidative stress. While high concentrations of H(2)O(2) caused an increase in protein sumoylation, low concentrations of H(2)O(2) mostly caused protein desumoylation. Immunofluorescence studies localized SUMO to the sites of DNA double-strand breaks in stressed germ cells and during meiotic recombination. To study the effect of oxidative stress in vivo, animals exposed to tobacco smoke for 12 weeks were used. Changes in sumoylation of HMW proteins were consistent with their oxidative damage in the tobacco-exposed mice. Our results are consistent with the important roles of different SUMO isoforms in stress responses in germ cells. Furthermore, this study identified topoisomerase 2 alpha as one of the targets of sumoylation during normal spermatogenesis and under stress.
Reproduction 04/2010; 139(6):999-1010. · 2.58 Impact Factor
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ABSTRACT: Small ubiquitin-like modifier (SUMO) proteins are involved in a variety of cellular processes. Alterations in SUMO conjugation have been implicated in several human diseases, including cancer. Although the main cause of failure in cancer treatment is the development of drug resistance by cancer cells, the mechanisms of drug resistance are not fully understood. SUMO proteins are thought to play roles in various cellular pathways, but no studies have as yet compared the expression of the different SUMO proteins in chemosensitive and drug-resistant cancer cells. To determine the relationship between protein sumoylation and drug resistance, the expression of various SUMO isoforms has been studied and compared in the HL-60 cell line (a model for leukemic cells) and in HL-60RV cells (resistant to vincristine). Co-immunostaining of cells by anti-SUMO antibodies and antibodies against various nuclear subdomains has been examined by an advanced type of bioimaging analysis. Whereas SUMO-2/3 co-localizes exclusively with nuclear bodies containing promyelocytic leukemia protein in both cell types, SUMO-1 has also been seen in nucleolar regions of HL-60, but not in HL-60RV, cells. In HL-60 cells, SUMO-1 occurs adjacent to, but not co-localized with, the nucleolar marker fibrillarin. Western blot analysis has revealed higher levels of free SUMO and sumoylated products in drug-resistant cells and the presence of specific SUMO-1 conjugates in drug-sensitive HL-60 cells, possibly consistent with a specific nucleolar signal. Shortly after the induction of ethanol and oxidative stress, HL-60RV, but not HL-60, cells show increased accumulation of high-molecular-weight SUMO-2/3 conjugates. Thus, SUMO-1 probably has a specific role in the nucleoli of HL-60 cells, and the alteration of sumoylation might be a contributing factor in the development of drug resistance in leukemia cells.
Cell and Tissue Research 04/2009; 336(2):277-86. · 3.11 Impact Factor
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Margarita Vigodner
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ABSTRACT: During meiosis in male mammals, X and Y chromosomes undergo the process of meiotic sex chromosome inactivation (MSCI). A crucial role in MSCI has recently been reported for BRCA1, ATR kinase, and phosphorylated histone H2AX, but the exact mechanism remains to be determined. Small ubiquitin-like modifier (SUMO) proteins have recently been shown to localize to the sex body in mouse meiotic spermatocytes, but the role they play during MSCI is unknown. In this study, in order to better understand the molecular events of MSCI, we followed dynamic changes in gammaH2AX and SUMO localization patterns during MSCI. Using confocal laser scanning microscopy (CLSM) as an analytical tool for visualizing numerous spermatocytes from the same development stage and for consecutively following the meiotic progression, we were able to demonstrate a very early appearance of SUMO-1, which preceded gammaH2AX accumulation on the sex chromosomes during their meiotic inactivation. In contrast to SUMO-1, SUMO-2/3 was undetectable in zygotene spermatocytes, suggesting a possible specific role for SUMO-1 in the initiation of MSCI.
Chromosome Research 02/2009; 17(1):37-45. · 3.09 Impact Factor
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ABSTRACT: Sumoylation affects multiple cellular events, including chromatin inactivation and transcriptional repression. Our data provide the first characterization of small ubiquitin-related modifier-1 (SUMO-1) expression during human spermatogenesis by the use of high-resolution cellular SUMO-1 bioimaging. During human meiotic prophase, SUMO-1 localizes to sex chromosomes and centromeric and pericentromeric chromatin. As human spermatocytes progress toward the end of prophase in meiosis I, SUMO-1 is no longer detected within the sex body and pericentromeric heterochromatin but localizes exclusively to centromeres. SUMO-1 localization along sex chromosome axes, pseudoautosomal region, and centromeres of both chromosomes supports a role for SUMO-1 sumoylation in epigenetic events occurring over the entire sex body, e.g., meiotic sex chromosome inactivation and chromatin condensation. Centromeric SUMO-1 throughout meiotic prophase suggests a role in centromeric chromatin condensation and/or other centromere/kinetochore functions. SUMO-1 is likely involved in both facultative and constitutive heterochromatin processes in spermatocytes. Haploid round spermatids show a consistent association of SUMO-1 with centromeric clusters. During spermatid elongation, SUMO-1 localizes in the manchette perinuclear ring. Steroidogenic Leydig cells show some cytoplasmic but strong nuclear and perinuclear SUMO-1. Peritubular myoepithelial cell SUMO-1 colocalizes with centromeric heterochromatin. In epithelial Sertoli cells, when associated with centromeric heterochromatin, SUMO-1 is adjacent but not colocalized with the nucleolus. Male germ cells demonstrate no SUMO-1 nucleolar association. Human and rodent Sertoli cells consistently show an inverse correlation between androgen receptor (AR) and SUMO-1 expression and compartmentalization. Sertoli cells from certain infertile patients, however, showed greatly decreased SUMO-1 and AR. Our data suggest that human testicular SUMO-1 has specific functions in heterochromatin organization, meiotic centromere function, and gene expression.
AJP Endocrinology and Metabolism 06/2006; 290(5):E1022-33. · 4.75 Impact Factor
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ABSTRACT: To compare the effect of two different techniques of testicular fixation on testicular function.
Experimental study.
Surgical animal laboratory at an academic medical center.
Sixteen mature golden hamsters underwent classic transfixation orchiopexy and true dartos pouch orchiopexy.
Classic transfixation orchiopexy (CTO) involved transfixation of the testicular wall at two different points and fixation of the dartos fascia. True dartos pouch orchiopexy (TDPO) involved creating a window in the dartos fascia, passage of the testicle, and closure of the window from both sides of the testicle.
Flow cytometric separation of testicular cells into haploid, diploid, and tetraploid fractions for histogram analysis.
A significant decrease in testicular weight was observed in 6 out of 16 animals undergoing CTO. Diploid cells comprised the main cell fraction, and almost no haploid or tetraploid cells were observed, while in the 16 animals undergoing TDPO no change from the control pattern was observed.
This experimental work supports our clinical impression that TDPO should replace CTO as the method of choice for the treatment of an undescended testicle in children.
Fertility and sterility 10/2005; 84(3):749-55. · 3.97 Impact Factor
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ABSTRACT: SUMO-1 is a member of a ubiquitin-related family of proteins that mediates important post-translational effects affecting diverse physiological functions. Whereas SUMO-1 is detected in the testis, little is known about its reproductive role in males. Herein, cell-specific SUMO-1 was localized in freshly isolated, purified male germ cells and somatic cells of mouse and rat testes using Western analysis, high-resolution single-cell bioimaging, and in situ confocal microscopy of seminiferous tubules. During germ cell development, SUMO-1 was observed at low but detectable levels in the cytoplasm of spermatogonia and early spermatocytes. SUMO-1 appeared on gonosomal chromatin during zygotene when chromosome homologues pair and sex chromatin condensation is initiated. Striking SUMO-1 increases in the sex body of early-to-mid-pachytene spermatocytes correlated with timing of additional sex chromosome condensation. Before the completion of the first meiotic division, SUMO-1 disappeared from the sex body when X and Y chromosomal activity resumed. Together, these data indicate that sumoylation may be involved in non-homologous chromosomal synapsis, meiotic sex chromosome inactivation, and XY body formation. During spermiogenesis, SUMO-1 localized in chromocenters of certain round spermatids and perinuclear ring and centrosomes of elongating spermatids, data implicating SUMO-1 in the process of microtubule nucleation and nuclear reshaping. STAT-4, one potential target of sumoylation, was located along the spermatid nuclei, adjacent but not co-localized with SUMO-1. Androgen receptor-positive Leydig, Sertoli, and some peritubular myoepithelial cells express SUMO-1, findings suggesting a role in modulating steroid action. Testicular SUMO-1 expression supports its specific functions in inactivation of sex chromosomes during meiosis, spermatid microtubule nucleation, nuclear reshaping, and gene expression.
Developmental Biology 07/2005; 282(2):480-92. · 4.07 Impact Factor
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ABSTRACT: Spermatogenesis consists of spermatogonial proliferation, meiosis and spermatid differentiation. Laser scanning confocal microscopy (LSCM) may be used as an advanced analytical tool to follow spermatogenesis inside the seminiferous tubules without performing histological sections. For this purpose, separated seminiferous tubules are fixed in 0.5% paraformaldehyde, stained for DNA with propidium iodide and analyzed by LSCM. By producing longitudinal optical sections in the layer of spermatogonia, spermatocytes and spermatids, stage-specific changes in their structure may be followed within the tubules by LSCM. Longitudinal z-sections may be obtained to produce three-dimensional images of the seminiferous tubules. In addition, different proteins may be followed during spermatogenesis in a stage specific manner within the tubule by incubation of the fixed seminiferous tubules with appropriate antibodies. As an example of the spermatogenesis studies using described LSCM techniques, detailed examination of spermatogonia, spermatocytes and spermatids during golden hamster spermatogenesis is presented. LSCM analysis of c-kit and SC3 protein expression at different stages of hamster spermatogenesis is demonstrated.
Methods in Cell Science 02/2002; 24(4):169-80.
