Analysis of the synaptonemal complex of the nine-banded armadillo, Dasypus novemcinctus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Conventional and ultrastructural analyses of the chromosomes of Discocyrtus pectinifemur (Opiliones, Laniatores, Gonyleptidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

SYNAPTONEMAL COMPLEX-ANALYSIS OF THE SEX BIVALENT IN GOATS

Synaptonemal complex (SC) analysis of XY pairing in the goat (Capra hircus; 2n = 60) was investigated by electron microscopy for the first time in this species. Synapsis of the X and Y chromosomes begins during the mid-late zygotene stage as the autosomes complete their pairing. Only a small portion of the total length of the Y is paired with the X chromosome at this time. By the early pachytene, almost 90% of the Y is paired with the X. All the observed stages of the sex bivalent pairing showed the structural difference between the differential and pairing regions. In the pairing region, a synaptonemal complex is formed, while in the differential region the chromosome axes remain free.

Reproductive compatibility and chromosome pairing in the Drosophila buzzatii complex

Fertility (percentage of fertile crosses) and the degree of synapsis in salivary gland chromosomes in isofemale lines of Drosophila buzzatii, D. serido, D. koepferae and D. seriema were analysed. D. buzzatii was completely sterile in intercrosses with strains from the other species except for D. koepferae. The other species intercrossed to a greater or lesser degree, but also differed between crossing directions. Homologous pairing in salivary gland preparations of strains and hybrids conformed with the data on fertility. The lowest degree of synapsis was present in hybrids between D. koepferae and D. buzzatii, D. seriema and D. koepferae and D. koepferae and D. serido. These species also exhibited the lowest degree of reproductive compatibility. Hybrids between D. seriema and D. serido showed an intermediate degree of synapsis (pairing absent in the proximal and distal chromosome ends), as well as fertility greater than that found in the other interspecific crosses. Results of the fertility of crosses involving strains of a single species, compared with data in the literature, indicated that intraspecific divergence occurred in D. serido and D. koepferae.

SYNAPTONEMAL COMPLEX-ANALYSIS IN SPERMATOCYTES OF TILAPIA, OREOCHROMIS-NILOTICUS (PISCES, CICHLIDAE)

Some adaptations of the synaptonemal complex (SC) whole-mounting technique first used in plants permitted its application to meiotic studies in tilapia, Oreochromis niloticus. Direct observation of the chromosome pairing process and bivalent structure during the meiotic prophase of this fish species by light and electron microscopy permitted the analysis of SCs in autosomes and the possible identification of sex chromosomes. The analysis of SCs in spermatocytes of 0. niloticus revealed that all 22 bivalent chromosomes completely paired, except for the occurrence of a size heteromorphism in the terminal region of the largest bivalent associated with the presence of an incompletely paired segment during the synapsis process, which may be the cytological visualization of an XX/XY sex chromosome system in this species.

Synaptonemal complex formation in spermatocytes of the autotriploid rainbow trout, Oncorhynchus mykiss (Pisces, Salmonidae)

Meiotic cells of triploid male rainbow trouts were analyzed by a surface-spreading SC technique in order to show the process of chromosome synapsis. At zygotene the formation of SCs involved, almost exclusively, two sets of lateral elements (LEs), and the remaining set of LEs presented different synaptic configurations involving one, two, three, or four LEs. The absence of SCs involving more than two LEs from mid- to late pachytene indicates that the multivalents produced by synapsis involving more than two LEs, are eliminated before the end of pachytene; thus, the mechanism of chromosome pairing in triploid male rainbow trouts produces, almost exclusively, bivalents with a probable extensive nonhomologous synapsis involving the extra set of chromosomes.

Paracentric inversion involving a NOR-bearing chromosome of rainbow trout (Oncorhyncus mykiss): Electron microscopy studies of the synaptonemal complex

Cytogenetic studies conduced on a sample of 19 specimens of rainbow trout (Oncorhyncus mykiss) showed that 6 presented two chromosomes with a single subterminal AgNOR on the short arms and 13 presented one chromosome with a single subterminal Ag-NOR and a second chromosome with two small subterminal Ag-NORs. Synaptonemal complex studies showed the presence of a pairing failure involving the two lateral elements (LEs) of the NOR-bearing chromosomes only in those specimens with two different NOR-bearing chromosomes, suggesting that the chromosomes with two Ag-NORs originated from a paracentric inversion involving a terminal segment of the original NOR-bearing chromosome. A comparative analysis between three normal and three heterozygous fishes suggest that in the latter group the LEs of the NOR-bearing chromosomes initiate the synaptic process in a more delayed manner, the synapsis develops more quickly, and the nucleolus stays associated with chromosomes for a longer time.

Cytogenetical and histological studies in testis of Tayassu tajacu (Cateto), Tayassu pecari (Queixada) and a natural interspecific hybrid

The karyotype, the histological structure of the testis and the synaptonemal complex (SC) of the mammalian species Tayassu tajacu, Tayassu pecari and of an interspecific male hybrid captured in nature were analysed. The specimens of T. tajacu (2n = 30) and T. pecan (2n = 26) exhibited seminiferous tubules with germ cells in all sperma to genesis stages. In the SC studies both species had a regular structure, easily identified in the autosomes and in the sex chromosomes. The hybrid (2n = 28) had seminiferous tubules with almost all germinal cells in the spermatogonium stage and only a few cells in the primary spermatocyte stage. Gross abnormalities in SC were observed. A few lateral elements showed regular or partially regular synapsis, other lateral elements were synapsed as multivalents, and most axial elements remained unsynapsed. The results suggest that the karyotypes of the parental species have sufficient differentiation to avoid chromosome synapsis. Alternatively, the hypothesis of the existence of genetic incompatibilities between the parental species is discussed.

Synaptonemal complex analysis in the fish species Piaractus mesopotamicus and Colossoma macropomum, and in their interspecific hybrid

The surface-spreading technique for visualization of whole cell synaptonemal complex (SC) complements was employed to study the chromosome synapsis in spermatocytes of P. mesopotamicus, C. macropomum and in their interspecific hybrid (tambacu) with the main objective to analyze possible errors in chromosome pairing that could result in hybrid sterility. SC analysis showed that the parental species P. mesopotamicus and C. macropomum have 27 bivalents homogeneously synapsed. The SC in spermatocytes from the hybrid tambacu showed gross meiotic configurations in all cells analyzed. The spermatocytes exhibited a few chromosomes or well synapsed chromosome segments, while many chromosome segments did not have any synapsis. This result, allied to other genetical and cytogenetical evidence, reinforces the hypothesis that the hybrid tambacu is sterile. Further studies involving other aspects, such as behavior and physiology, should be conduced before the introduction of these hybrids in rivers and lakes.

Chromosomal similarities and differences among four Neotropical Elateridae (Conoderini and Pyrophorini) and other related species, with comments on the NOR patterns in Coleoptera

This work deals with the comparative cytogenetic analysis of four Neotropical Elateridae species and reviews the nucleolar organizer region (NOR) patterns on Coleoptera chromosomes, for the first time. The cytogenetic characterization of Conoderus malleatus (Conoderini), Pyrearinus candelarius, Pyrophorus divergens and Pyrophorus punctatissimus (Pyrophorini) was accomplished through the study of mitotic and meiotic cells submitted to standard (Giemsa) and differential staining [silver impregnation and GC-specific chromomycin A 3 (CMA 3) plus AT-specific 4′-6-diamidino-2-phenylindole (DAPI) fluorochromes]. The analysis of spermatogonial cells revealed the diploid numbers: 2n = 17 in C. malleatus and 2n = 15 in P. candelarius, P. divergens and P. punctatissimus. In these species, the X0 type sex-determination system and the acrocentric morphology of almost all chromosomes were observed. The study of meiotic cells of the four species revealed the occurrence of total synapsis between the autosomes, the presence of one terminal or interstitial chiasma in the majority of the bivalents, and the reductional behaviour and regular segregation of all chromosomes. Although the three Pyrophorini species demonstrated many similar karyotypical characteristics, there was one discrepancy, which was noted in the diplotene cells and concerns the number of bivalents with two chiasmata; P. candelarius only presented one bivalent, P. divergens showed two bivalents and P. punctatissimus exhibited up to four bivalents with two chiasmata. Testicular cells impregnated with silver nitrate demonstrated two terminal NORs located on the fourth autosomal pair of the Conoderini species and on the second autosomal pair of the three Pyrophorini representatives. Use of CMA 3/distamycin A (DA)/DAPI staining on the P. candelarius and P. punctatissimus chromosomes revealed that the CMA 3 labelled regions were coincident with the NORs. The main strategies of karyotypical differentiation that have occurred among the four Elateridae species and other related species, and the general trends of the NOR shifts during Coleoptera chromosomal evolution are discussed in this work. © 2007 The Authors.

A NEURAL NET FOR EXTRACTING KNOWLEDGE FROM NATURAL-LANGUAGE DATA-BASES

The present paper introduces a new model of fuzzy neuron, one which increases the computational power of the artificial neuron, turning it also into a symbolic processing device. This model proposes the synapsis to be symbolically and numerically defined, by means of the assignment of tokens to the presynaptic and postsynaptic neurons. The matching or concatenation compatibility between these tokens is used to decided about the possible connections among neurons of a given net. The strength of the compatible synapsis is made dependent on the amount of the available presynaptic and post synaptic tokens. The symbolic and numeric processing capacity of the new fuzzy neuron is used here to build a neural net (JARGON) to disclose the existing knowledge in natural language data bases such as medical files, set of interviews, and reports about engineering operations.

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.

Interactions of RecF protein with RecO, RecR, and single-stranded DNA binding proteins reveal roles for the RecF–RecO–RecR complex in DNA repair and recombination

The products of the recF, recO, and recR genes are thought to interact and assist RecA in the utilization of single-stranded DNA precomplexed with single-stranded DNA binding protein (Ssb) during synapsis. Using immunoprecipitation, size-exclusion chromatography, and Ssb protein affinity chromatography in the absence of any nucleotide cofactors, we have obtained the following results: (i) RecF interacts with RecO, (ii) RecF interacts with RecR in the presence of RecO to form a complex consisting of RecF, RecO, and RecR (RecF–RecO–RecR); (iii) RecF interacts with Ssb protein in the presence of RecO. These data suggested that RecO mediates the interactions of RecF protein with RecR and with Ssb proteins. Incubation of RecF, RecO, RecR, and Ssb proteins resulted in the formation of RecF–RecO–Ssb complexes; i.e., RecR was excluded. Preincubation of RecF, RecO, and RecR proteins prior to addition of Ssb protein resulted in the formation of complexes consisting of RecF, RecO, RecR, and Ssb proteins. These data suggest that one role of RecF is to stabilize the interaction of RecR with RecO in the presence of Ssb protein. Finally, we found that interactions of RecF with RecO are lost in the presence of ATP. We discuss these results to explain how the RecF–RecO–RecR complex functions as an anti-Ssb factor.

Replication-dependent early meiotic requirement for Spo11 and Rad50

Spo11 and the Rad50-Mre11 complex have been indirectly implicated in processes associated with DNA replication. These proteins also have been shown to have early meiotic roles essential for the formation of a programmed DNA double-strand break known in Saccharomyces cerevisiae to initiate meiotic recombination. In both S. cerevisiae and the basidiomycete Coprinus cinereus, spo11 and rad50 mutants are defective in chromosome synapsis during meiosis. Here we demonstrate that a partial restoration of synapsis occurs in C. cinereus spo11 and rad50 mutants if premeiotic DNA replication is prevented. Double mutants were constructed with spo11–1 or rad50–4 and another mutant, spo22–1, which does not undergo premeiotic DNA replication. In both cases, we observed an increase in the percentage of nuclei containing synaptonemal complex (SC) structures, with concomitant decreases in the percentage of nuclei containing axial elements (AE) only or no structures. Both types of double mutants demonstrated significant increases in the average numbers of AE and SC, although SC-containing nuclei did not on average contain more AE than did nuclei showing no synapsis. Our results show that Spo11-induced recombination is not absolutely required for synapsis in C. cinereus, and that the early meiotic role of both Spo11 and Rad50 in SC formation partially depends on premeiotic S phase. This dependency likely reflects either a requirement for these proteins imposed by the premeiotic replication process itself or a requirement for these proteins in synapsis when a sister chromatid (the outcome of DNA replication) is present.

Nucleotide excision repair in rat male germ cells: low level of repair in intact cells contrasts with high dual incision activity in vitro

The acquisition of genotoxin-induced mutations in the mammalian germline is detrimental to the stable transfer of genomic information. In somatic cells, nucleotide excision repair (NER) is a major pathway to counteract the mutagenic effects of DNA damage. Two NER subpathways have been identified, global genome repair (GGR) and transcription-coupled repair (TCR). In contrast to somatic cells, little is known regarding the expression of these pathways in germ cells. To address this basic question, we have studied NER in rat spermatogenic cells in crude cell suspension, in enriched cell stages and within seminiferous tubules after exposure to UV or N-acetoxy-2-acetylaminofluorene. Surprisingly, repair in spermatogenic cells was inefficient in the genome overall and in transcriptionally active genes indicating non-functional GGR and TCR. In contrast, extracts from early/mid pachytene cells displayed dual incision activity in vitro as high as extracts from somatic cells, demonstrating that the proteins involved in incision are present and functional in premeiotic cells. However, incision activities of extracts from diplotene cells and round spermatids were low, indicating a stage-dependent expression of incision activity. We hypothesize that sequestering of NER proteins by mispaired regions in DNA involved in synapsis and recombination may underlie the lack of NER activity in premeiotic cells.