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Biomedical subjects

R Benavente

Publications and source records attributed to R Benavente.

At least 19 recordsLinked to original sources
PubMed2003

Microprolactinomas in males treated by transsphenoidal surgery.

OBJECTIVE: To support the opinion that transsphenoidal surgery can be an effective alternative to medical treatment for microprolactinomas in men. DESIGN: Clinical study with retrospective data analysis. PATIENTS AND METHODS: Of 46 men who were operated on for prolactinoma in the Department of Neurosurgery of the University of Vienna General Hospital between 1985 and 2000 a microadenoma was detected 11 times (24%). RESULTS: Median patient age was 41 years (range 32 to 54 years). Symptoms were of endocrine nature in all patients with erectile dysfunction, infertility and gynaecomastia being the initial complaints and having lasted for a median of 13 months (range 7-68 months). Preoperative median serum prolactin (PRL) was elevated to 120 ng/ml (range 41-1000 ng/ml). Radiography by MRI revealed microadenomas with a median diameter of 8 mm (range 4-10 mm). All patients were operated on via the transsphenoidal approach. Endocrine cure as defined by a serum PRL <25 ng/ml was achieved in 8 of 11 patients (73%) after a median follow-up of 7 years (range 2-13 years). In none of the 3 patients with preoperative serum PRL levels >150 ng/ml was a normoprolactinaemia obtained after surgery: 2 require further dopamine-agonist therapy after surgery, 1 after late follow-up. Surgery and medical treatment could restore potency and libido in all but 1 patient, 2 of 3 patients remain infertile. CONCLUSION: Prolactinomas in males are potentially curable by surgery if detected at an early stage. They most commonly present with insidious signs and symptoms of endocrine disturbances such as loss of libido, impotence and sterility. We stress the importance of early determination of serum prolactin and high resolution magnetic resonance tomography of the sella in male patients with hyperprolactinaemia as this may prevent the possible progression to larger tumours which are rarely curable by surgery and necessitate life-long medical therapy.

Adult↗
PubMed2003

Histone H1t is not replaced by H1.1 or H1.2 in pachytene spermatocytes or spermatids of H1t-deficient mice.

The linker histone gene H1t is exclusively expressed in the mammalian testis. In former experiments we have shown that H1.1 and H1.2 histone gene expression is significantly enhanced in testis of adult H1t deficient mice. In this report we have quantified the mRNA of different H1 genes in 9-day- and 20-day-old wild type and H1t knock out mice. In addition, we have analysed the distribution of H1.1 and H1.2 protein by immunofluorescent staining in spread male germ cells. The aim of this work was to answer the question whether H1t can be replaced during spermatogenesis by H1.1 or H1.2. In our experiments we could not detect elevated levels of H1.1 or H1.2 in pachytene spermatocytes or haploid cells of H1t deficient testis. Therefore, in these cells, H1t seems not to be replaced by H1.1 or H1.2.

Animals↗
PubMed2003

Ultrastructural and immunocytochemical analysis of the XY body in rat and Guinea pig.

The formation of the XY body involves the compaction of the extended chromatin to form a mesh of fibrogranular structures. During this process the ribonucleoprotein particles (RNP), which were associated with the chromatin filaments progressively disappear. High resolution immunolocalization indicates that the mature XY body does not contain RNA polymerase II, hnRNPs, or snURNPs. Occasionally chromatin fibrils extend outside of the XY body. These fibrils are frequently associated with nascent RNP fibrils and granules indicating that not all the DNA of the sex chromosomes is transcriptionally inactive. However, transcription is located outside the sex body. The recombination protein Dmc1 is present in nodules associated with the unpaired chromosomal axes of the sex chromosomes located in the XY body. Cytochemical staining methods and in situ hybridization at electron microscopic level show that RNA is present in the unpaired chromosomal axes suggesting that the presence of RNA in the chromosomal axes and in forming synaptonemal complexes is related with the process of final pairing. The sex body and the nucleoli associated with it do not interweave and do not exchange RNA or DNA-containing filaments. These observations indicate that the spatial relation between these structures is just a close proximity, which is, however, very frequent.

Animals↗
PubMed2002

Identification and characterization of SRSP1, a rat gene differentially expressed during spermatogenesis and coding for a serine stretch-containing protein.

By means of mRNA differential display and cDNA library screening we have characterized a novel gene of the rat that is differentially expressed during spermatogenesis. Northern blot analyses showed that its mRNA reaches the highest level during the first meiotic prophase. The transcript appears to be testis-specific as it was not detectable in any of the nine other tissues tested. The full length ORF encodes a putative phosphoprotein containing a serine stretch and a bipartite nuclear localization signal, which we call Srsp1 ("serin-rich spermatogenic protein 1"). Comparison of the cDNA from SRSP1 with genomic sequences in databases allowed us to determine the number of introns; some putative regulatory sequences are proposed. This is the first report of a gene differentially expressed during spermatogenesis that codes for a protein containing a serine stretch.

Amino Acid Sequence↗
PubMed2000

Meiotic lamin C2: the unique amino-terminal hexapeptide GNAEGR is essential for nuclear envelope association.

Meiotic lamin C2 is the only A-type lamin expressed during mammalian spermatogenesis. Typical for this short lamin is the unique hexapeptide GNAEGR, which substitutes the nonhelical amino terminus and part of the alpha-helical rod domain present in somatic lamins. Meiotic lamin C2 also lacks a carboxyl-terminal CaaX box, which is modified by isoprenylation and involved in nuclear envelope (NE) association of somatic isoforms. The mechanism by which lamin C2 becomes localized in the NE is totally unknown. Here we demonstrate that the hexapeptide GNAEGR is essential for this process: (i) Its deletion resulted in a diffuse distribution of lamin C2 within nuclei of transfected COS-7 cells; (ii) Mutated somatic lamin C, containing the sequence GNAEGR at its amino terminus, was located at the NE. The mass spectrometric analysis of the amino terminus of lamin C2 revealed that it is modified by myristoylation. Correspondingly, the substitution of the first glycine residue abolishes the NE association of lamin C2. We conclude that NE association of lamin C2 is achieved by a mechanism different from that of somatic lamins.

Amino Acid Sequence↗
PubMed2000

Analysis of male meiotic "sex body" proteins during XY female meiosis provides new insights into their functions.

During male meiosis in mammals the X and Y chromosomes become condensed to form the sex body (XY body), which is the morphological manifestation of the process of meiotic sex chromosome inactivation (MSCI). An increasing number of sex body located proteins are being identified, but their functions in relation to MSCI are unclear. Here we demonstrate that assaying male sex body located proteins during XY female mouse meiosis, where MSCI does not take place, is one way in which to begin to discriminate between potential functions. We show that a newly identified protein, "Asynaptin" (ASY), detected in male meiosis exclusively in association with the X and Y chromatin of the sex body, is also expressed in pachytene oocytes of XY females where it coats the chromatin of the asynapsed X in the absence of MSCI. Furthermore, in pachytene oocytes of females carrying a reciprocal autosomal translocation, ASY associates with asynapsed autosomal chromatin. Thus the location of ASY to the sex body during male meiosis is likely to be a response to the asynapsis of the non-homologous regions [outside the pseudoautosomal region (PAR)] of the heteromorphic X-Y bivalent, rather than being related to MSCI. In contrast to ASY, the previously described sex body protein XY77 proved to be male sex body specific. Potential functions for MSCI and the sex body are discussed together with the possible roles of these two proteins.

Animals↗
PubMed2000

Fate of meiotic lamin C2 in rat spermatocytes cultured in the presence of okadaic acid.

The fate of nuclear envelope proteins during the pachytene/metaphase I transition was investigated in rat spermatocytes cultured in vitro in the presence of the phosphatase inhibitor okadaic acid (OA). Under these experimental conditions lamin B1 and the lamina-associated proteins 2 (LAPs2) behave as already described in other cell types. In contrast to these results, meiotic lamin C2 appears to be degraded after addition of OA to the spermatocyte culture medium as this lamin was no longer detectable by immunofluorescence microscopy or by immunoblotting. Taking into account the peculiarities of the lamin C2 primary structure, it is tempting to speculate that degradation of this protein represents a critical step in the process of disassembly of the spermatocyte nuclear envelope.

Animals↗
PubMed1999

Architecture of the nuclear periphery of rat pachytene spermatocytes: distribution of nuclear envelope proteins in relation to synaptonemal complex attachment sites.

The nucleus of spermatocytes provides during the first meiotic prophase an interesting model for investigating relationships of the nuclear envelope (NE) with components of the nuclear interior. During the pachytene stage, meiotic chromosomes are synapsed via synaptonemal complexes (SCs) and attached through both ends to the nuclear periphery. This association is dynamic because chromosomes move during the process of synapsis and desynapsis that takes place during meiotic prophase. The NE of spermatocytes possesses some peculiarities (e.g., lower stability than in somatic cells, expression of short meiosis-specific lamin isoforms called C2 and B3) that could be critically involved in this process. For better understanding of the association of chromosomes with the nuclear periphery, in the present study we have investigated the distribution of NE proteins in relation to SC attachment sites. A major outcome was the finding that lamin C2 is distributed in the form of discontinuous domains at the NE of spermatocytes and that SC attachment sites are embedded in these domains. Lamin C2 appears to form part of larger structures as suggested by cell fractionation experiments. According to these results, we propose that the C2-containing domains represent local reinforcements of the NE that are involved in the proper attachment of SCs.

Animals↗
PubMed1999

Molecular characterization and developmentally regulated expression of Xenopus lamina-associated polypeptide 2 (XLAP2).

Lamina-associated polypeptides 2 (LAP2alpha, beta, gamma)/thymopoietins (TPalpha, beta, gamma) are a family of proteins that are generated by alternative splicing from a single gene. These proteins have been primarily characterized in mammals. One member of this protein family, the integral membrane protein LAP2beta/TPbeta, has been localized to the inner nuclear membrane of somatic cells where it binds to chromatin and B-type lamins. By cDNA cloning we have characterized XLAP2, a Xenopus homologue of the mammalian LAP2beta. Using LAP2-specific antibodies, the Mr 68,000 XLAP2 was found to be the only member of the LAP2/TP family expressed in somatic cells and adult tissues. XLAP2 was not detected in oocytes, eggs and in early embryos up to the gastrula stage at the mRNA and protein level demonstrating that it is not synthesized from maternal mRNA. In counterpart oocytes, eggs, and embryos contained one LAP2-related integral membrane proteins of Mr 84,000. Northern blot analysis with the XLAP2 cDNA showed that a single hybridizing mRNA band of 1.8-2.0 kb was present in Xenopus somatic cells whereas two other hybridizing mRNA species of 2.8-3.0 and 0. 9-1.1 kb were present in oocytes, eggs and early embryos. All together, these results indicated that at least three distinct LAP2-related proteins might be expressed in Xenopus. The LAP2/TP protein of Mr 84,000 is present in the early embryos but its amount decreases during embryogenesis concomitant with the increase of XLAP2 in the embryo. Our results are the first description of the developmentally regulated expression of integral nuclear envelope proteins during early embryogenesis.

Amino Acid Sequence↗
PubMed1998

The murine nuclear orphan receptor GCNF is expressed in the XY body of primary spermatocytes.

We have studied the expression of the nuclear orphan receptor GCNF (germ cell nuclear factor) on the mRNA and protein level in pubertal and adult mouse testes. We show by Northern and Western blot analyses and by in situ hybridization that GCNF is expressed in spermatocytes and round spermatids of adult mouse testis suggesting that GCNF may be a transcriptional regulator of spermatogenesis. Since the GCNF protein is accumulated in the XY body of late pachytene spermatocytes, it may be involved in transcriptional inactivation of sex chromosomes.

Animals↗
PubMed1998

Nuclear envelope remodelling during rat spermiogenesis: distribution and expression pattern of LAP2/thymopoietins.

Lamina-associated polypeptide 2 (LAP2) and the thymopoietins (TPs) are a family of proteins described in somatic cells of mammals, which are derived by alternative splicing from a single gene. For one of the members of the family (LAP2 = TPbeta) it has been shown that this integral membrane protein locates to the inner membrane of the nuclear envelope, and that it binds to chromatin and B-type lamins. In the present study, we observed that during the third phase of spermatogenesis (i.e. spermiogenesis), TP-labelling shifted progressively to one half of the nuclear periphery in round spermatids. In the elongating spermatid the signal then becomes restricted to one spot located at the posterior (centriolar) pole of the nucleus. Changes in localization are accompanied by the disappearance, first of TPgamma, and later on of LAP2/TPbeta. TPalpha is the only member of the family detectable in the mature sperm. Concomitantly, lamin B1, the only nuclear lamina protein known to be expressed in mammalian spermatids, showed a similar behaviour, i.e. shifted progressively to the centriolar pole of spermatid nuclei before it became undetectable in fully differentiated mature sperms. These results are the first demonstration that expression and localization patterns of TPs are coordinately and differentially regulated with lamins during a differentiation process.

Animals↗
PubMed1997

A novel Mr 77,000 protein of the XY body of mammalian spermatocytes: its localization in normal animals and in Searle's translocation carriers.

We describe a novel XY body protein of rat and mice pachytene spermatocytes called XY77. Biochemical characterization showed that protein XY77 (Mr 77,000; pH value 8.3) is present in meiotic but absent in postmeiotic stages of spermatogenesis. With the aid of an antibody against protein XY77 together with another specific for XY body-associated protein XY40 we also investigated the localization of these proteins in mice carrying Searle's translocation, a reciprocal X-autosomal translocation. We show here that in these mice the distribution of both XY77 and XY40 is abnormal. Our results indicate that in Searle's translocation alterations are not restricted to the translocated autosome, but also involve chromatin segments corresponding originally to the sex chromosomes X and Y.

Animals↗
PubMed1997

XY body formation during rat spermatogenesis: an immunocytochemical study using antibodies against XY body-associated proteins.

The process of formation of the XY body during meiotic prophase was investigated by immunocytochemistry on cryosections of pubertal rat testes using antibodies against three different XY body-associated proteins. Here we show that these proteins are detectable at only partially overlapping temporal windows. These findings provide the first evidence that the previously described morphological changes in the structure of the XY body that occur during meiotic prophase are accompanied by considerable changes in its protein composition.

Animals↗
PubMed1997

Molecular characterization and expression pattern of XY body-associated protein XY40 of the rat.

The XY body is a structure formed by the partially synapsed chromosomes X and Y that is located at the nuclear periphery of mammalian pachytene spermatocytes. In contrast to the autosomal bivalents of the same nucleus, the XY body is characterized by its differential chromatin condensation and transcriptional inactivity. In order to shed some light on the biological significance of these differences we have been characterizing XY body-associated proteins. We present here the cDNA sequence and expression pattern of XY40, a protein that is associated with the axial elements of the XY body. RNA blot analysis revealed that during spermatogenesis the transcript that encodes protein XY40 was highly enriched in pachytene spermatocytes. This transcript was also detectable in brain and, to a lesser extent, in liver and kidney. Although the signal in brain was as strong as in spermatocytes, protein XY40 could be detected only in the latter. The nucleic acid sequence reveals that XY40 is a novel protein with a few similarities to already known nucleotide sequences. Among these similarities the most interesting is a box that is shared by the 3' untranslated region of XY40 and the 5' untranslated region of Munc-18c, a member of a protein family involved in synaptic vesicle exocytosis. Since the transcripts of both XY40 and Munc18-c show a similar expression pattern, it is tempting to speculate that this common sequence is involved in translation regulation.

Amino Acid Sequence↗
PubMed1996

Change of karyoskeleton during mammalian spermatogenesis: expression pattern of nuclear lamin C2 and its regulation.

Nuclear lamins are a multigene family of major karyoskeletal proteins. The expression pattern of members of the lamin family has been shown to be developmentally regulated. Of particular interest have been the findings that mammalian spermatogenic cells express two small lamin isoforms (B3 and C2) which are shorter splicing variants of their somatic counterparts (the lamins B2 and C, respectively). Although lamins B3 and C2 appear to be specific for the germ line, the expression pattern and localization of these proteins is not known. In the present study, we present evidence that during rat spermatogenesis lamin C2 is selectively expressed in spermatocytes, i.e., the cells undergoing meiosis. As in the case of other members of the family, lamin C2 was detected at the nuclear periphery. Northern blotting and in situ hybridization indicate that meiotic expression of lamin C2 is regulated at the transcriptional level. This is in contrast to the situation during amphibian oogenesis, where lamin expression is largely posttrascriptionally regulated. Interestingly, the expression pattern of lamin C2 temporally coincides with that of structural protein components of another meiosis-specific karyoskeletal structure, the synaptonemal complex (SC). Taken together, we conclude that pairing and recombination of homologous chromosomes during meiotic prophase is accompanied by significant changes in the organization of the karyoskeleton which are accomplished by the expression of stage-specific proteins.

Amino Acid Sequence↗
PubMed1996

The mouse poly(C)-binding protein exists in multiple isoforms and interacts with several RNA-binding proteins.

The murine poly(C)-binding protein (mCBP) was previously shown to belong to the group of K-homology (KH) proteins by virtue of its homology to hnRNP-K. We have isolated cDNA-splice variants of mCBP which differ by two variable regions of 93 bp and/or 39 +/- 3 bp respectively. Both variable regions are located between the second and third KH-domain of mCBP. The characterization of a partial genomic clone enabled us to propose a model for the generation of the second variable region by the use of a putative alternative splice signal. The mCBP mRNA is expressed ubiquitously and the protein is found predominantly in the nucleus with the exception of the nucleoli. We have identified five proteins which interact with mCBP in the yeast two hybrid system: mouse y-box protein 1 (msy-1), y-box-binding protein, hnRNP-L, filamin and splicing factor 9G8. The interaction between mCBP and splicing factor 9G8 was confirmed in vivo. These results suggest a function of mCBP in RNA metabolism.

Animals↗