Light microscope analysis of meiotic prophase chromosomes by silver staining

Fletcher, J.

doi:10.1007/bf00293237

Cited by 127 publications

(41 citation statements)

References 18 publications

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“…1) Cells were suspended within drops of RNA-preserving buffer CSK [10 mM piperazine-N,N-bis(2-ethanesulfonic acid (PIPES), pH 6.8, 100 mM NaCl, 300 mM sucrose, 3 mM MgCl2 (Fey et al, 1986)] placed on silane coated slides, permeabilized on ice with Triton X-100 in CSK for 3 min, washed and fixed in 3.7% paraformaldehyde in CSK and then immediately immunostained. 2) Cells were microspread on clean slides in drops of 0.2 M sucrose, fixed in 4% paraformaldehyde in 0.1 M sucrose adjusted to pH 8.5, and washed in Photoflo (Fletcher, 1979). Slides were either stained with 50% silver nitrate for visualization of autosomal synaptonemal complexes and sex chromosome axes, or stored at -80°C until used.…”

Section: Cell Preparationmentioning

confidence: 99%

Splicing components are excluded from the transcriptionally inactive XY body in male meiotic nuclei.

Richler

Ast

Goitein

et al. 1994

MBoC

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The study of the effect of programmed cessation of transcription in a large nuclear domain, on the distribution of elements of the pre-mRNA splicing machinery, is the main aim of this paper. To this end, we took advantage of the nuclear partitioning of mouse spermatocytes early in meiosis into autosomal transcribing and XY nontranscribing compartments. This system also allows to extend this study to stages in sperm differentiation that are accompanied by reduction and eventual cessation of transcription. We show by indirect immunofluorescence in spermatogenetic cells that 1) fluorescent signals of the pre-mRNA splicing factors SF53/4 and SC35, of the Sm antigens, and of RNA polymerase II, are largely absent from the nontranscribing, X-inactivated compartment, but are abundantly present in the transcribing autosomal compartment and 2) the presence, gradual reduction, and absence of transcriptive activity in nuclei undergoing the sperm formation sequence are positively correlated with the fluorescence patterns of the antibodies against SF53/4, SC35, and the Sm antigens. These data suggest that cessation of transcription during spermatogenesis is accompanied by exclusion of the splicing machinery from nontranscribing chromatin to its vicinity. INTRODUCTIONThe perfection of the nucleus leading to the evolution of the eukaryotic cell was achieved by structural adaptations to the requirements of DNA replication, transcription, RNA processing, and message transportation. This paper deals primarily with one aspect of nuclear functional organization, namely, the effect of cessation of transcription in a large nuclear region located within a transcribing environment on the distribution of premRNA splicing components.The maturation of most eukaryotic pre-mRNA involves posttranscriptional processing events that are required to convert the nuclear primary transcripts of RNA

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Section: Cell Preparationmentioning

confidence: 99%

Splicing components are excluded from the transcriptionally inactive XY body in male meiotic nuclei.

Richler

Ast

Goitein

et al. 1994

MBoC

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“…The preparation was centrifuged and the cells were resuspended in a small quantity of the medium. The suspension was dropped onto a solution of 0.5M sucrose (FLETCHER 1979) on a plastic coated slide (FELLUGA and MARTINUCCI 1976;DRESSE-.R and MOSFS 1979). The cells were allowed to settle for about ten min.…”

Section: Methodsmentioning

confidence: 99%

The behavior during pachynema of a normal and an inverted Y chromosome in Microtus agrestis

2008

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The pachytene behavior of the chromosomes of Microrus agresris (L.) (Rodentia, Arvicolidae) males carrying either the standard, or the pencentrically inverted Lund Y chromosome have been examined by electron microscopy of microspread spermatocytes. There is no synapsis between the X and either the standard or the Lund Y chromosomes during any substage of pachynema. Since synapsis is generally considered a prerequisite for crossing over, there appears to be no opportunity for crossover or chiasma formation between the X and Y in this species. The G-, C-and NOR-banded mitotic karyotypes of animals carrying the standard and Lund Y are also presented. KOLLER and DARLINGTON (1934) observed a region of synapsis between the X and Y chromosomes of the Norwegian rat during pachynema and end association between them during metaphase I of meiosis. Since it was (and is) assumed that the mammalian X and Y arose from an ancestral pair of homologues, they concluded that there is a region of homology between the X and Y in all mammalian species and that this region regularly synapses and crosses over during meiosis in a manner analogous to the autosomes. This assumption is still widely accepted (BURGOYNE 1982;POLANI 1982). Indeed, the idea has gained recent momentum from the molecular data for regular exchange of DNA between the X and Y in humans (ROUYER et al. 1986). None the less, there is ample evidence that synapsis and subsequent crossover of the X and Y as a mechanism of disjunction of the sex chromosomes is not a universal phenomenon in mammals, or even in eutherians (cf. ASHLEY 1985ASHLEY , 1987 for review).One species which does not fit this general pattern is the Egyptian sand rat (Psammomys obesus), in which there is clearly no synapsis between the X and Y during pachynema (SOLARI and ASHLEY 1977;ASHLEY and MOSES 1980). Since synapsis is generally assumed to be a prerequisite for crossing over, it can be presumed that the asynapsis of the X and Y provides no opportunity for crossover. Consequently, disjunction of the sex chromosomes must be assumed to be achiasmatic in this species. The most prominent feature of the karyotype of the sand rat is formed by large blocks of heterochromatin on both the X and Y. SOLARI and ASHLEY (1977) suggested that this heterochromatin might be involved in an unspecified manner in the asynaptic behavior of the sex chromosomes of this species. It was therefore of interest to determine whether other species with large blocks of heterochromatin on their sex chromosomes are also asynaptic, or whether they synapse with a potential for chiasmatic disjunction. Microtus agrestis, the European field vole, has the distinction of having more constitutive heterochromatin on its sex chromosomes than any other species so far examined. Heterochromatin accounts for approximately 25 YO of the total female haploid genome (Hsu and ARRIGHI 1971;GAMPERL et al. 1982) of which 20 Yo is located on the X chromosome. Approximately three fourths of the submetacentric X is heterochromatic; the rest is euchromati...

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“…Surface spreading of meiocytes has provided a good means for the analysis of synaptonemal complexes (SCs) in animal material (CouNCE and MEYER 1973;FLETCHER 1979;DRESSER and MosEs 1980). A nearly identical method was successfully applied to maize microsporocytes by GILLIES (1981GILLIES ( , 1983.…”

Section: Introductionmentioning

confidence: 99%

Light Microscopical Observations on Surface Spread Synaptonemal Complexes ofAllium Ursinum

Loidl

1984

Caryologia

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Light microscope analysis of meiotic prophase chromosomes by silver staining

Cited by 127 publications

References 18 publications

Splicing components are excluded from the transcriptionally inactive XY body in male meiotic nuclei.

Splicing components are excluded from the transcriptionally inactive XY body in male meiotic nuclei.

The behavior during pachynema of a normal and an inverted Y chromosome in Microtus agrestis

Light Microscopical Observations on Surface Spread Synaptonemal Complexes ofAllium Ursinum

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