Chromosomal organization of Xenopus laevis oocyte and somatic 5S rRNA genes in vivo

1992 ◽  
Vol 12 (1) ◽  
pp. 45-55
Author(s):  
C C Chipev ◽  
A P Wolffe
Keyword(s):  
Xenopus Laevis ◽  
Rrna Genes ◽  
Dna Repeats ◽  
5S Rna ◽  
Chromosomal Organization ◽  
Dna Repeat ◽  
5S Dna ◽  
Nuclease Digestion ◽  
5S Rna Genes ◽  
Rna Genes

We describe the chromosomal organization of the major oocyte and somatic 5S RNA genes of Xenopus laevis in chromatin isolated from erythrocyte nuclei. Both major oocyte and somatic 5S DNA repeats are associated with nucleosomes; however, differences exist in the organization of chromatin over the oocyte and somatic 5S RNA genes. The repressed oocyte 5S RNA gene is protected from nuclease digestion by incorporation into a nucleosome, and the entire oocyte 5S DNA repeat is assembled into a loosely positioned array of nucleosomes. In contrast, the potentially active somatic 5S RNA gene is accessible to nuclease digestion, and the majority of somatic 5S RNA genes appear not to be incorporated into positioned nucleosomes. Evidence is presented supporting the stable association of transcription factors with the somatic 5S RNA genes. Histone H1 is shown to have a role both in determining the organization of nucleosomes over the oocyte 5S DNA repeat and in repressing transcription of the oocyte 5S RNA genes.

scholarly journals Chromosomal organization of Xenopus laevis oocyte and somatic 5S rRNA genes in vivo.

1992 ◽  
Vol 12 (1) ◽  
pp. 45-55 ◽  
Author(s):  
C C Chipev ◽  
A P Wolffe
Keyword(s):  
Xenopus Laevis ◽  
Rrna Genes ◽  
Dna Repeats ◽  
5S Rna ◽  
Chromosomal Organization ◽  
Dna Repeat ◽  
5S Dna ◽  
Nuclease Digestion ◽  
5S Rna Genes ◽  
Rna Genes

We describe the chromosomal organization of the major oocyte and somatic 5S RNA genes of Xenopus laevis in chromatin isolated from erythrocyte nuclei. Both major oocyte and somatic 5S DNA repeats are associated with nucleosomes; however, differences exist in the organization of chromatin over the oocyte and somatic 5S RNA genes. The repressed oocyte 5S RNA gene is protected from nuclease digestion by incorporation into a nucleosome, and the entire oocyte 5S DNA repeat is assembled into a loosely positioned array of nucleosomes. In contrast, the potentially active somatic 5S RNA gene is accessible to nuclease digestion, and the majority of somatic 5S RNA genes appear not to be incorporated into positioned nucleosomes. Evidence is presented supporting the stable association of transcription factors with the somatic 5S RNA genes. Histone H1 is shown to have a role both in determining the organization of nucleosomes over the oocyte 5S DNA repeat and in repressing transcription of the oocyte 5S RNA genes.

Regular arrangement of nucleosomes on 5S rRNA genes in Xenopus laevis

1983 ◽  
Vol 3 (4) ◽  
pp. 720-730
Author(s):  
D Young ◽  
D Carroll
Keyword(s):  
Xenopus Laevis ◽  
Chromatin Structure ◽  
Micrococcal Nuclease ◽  
Rrna Genes ◽  
Dna Repeats ◽  
5S Rna ◽  
5S Dna ◽  
Regular Arrangement ◽  
5S Rna Genes ◽  
Rna Genes

The chromatin structure of the oocyte-type 5S RNA genes in Xenopus laevis was investigated. Blot hybridization analysis of DNA from micrococcal nuclease digests of erythrocyte nuclei showed that 5S DNA has the same average nucleosome repeat length, 192 +/- 4 base pairs, as two Xenopus satellite DNAs and bulk erythrocyte chromatin. The positions of nuclease-sensitive regions in the 5S DNA repeats of purified DNA and chromatin from erythrocytes were mapped by using an indirect end-labeling technique. Although most of the sites cleaved in purified DNA were also cleaved in chromatin, the patterns of intensities were strikingly different in the two cases. In 5S chromatin, three nuclease-sensitive regions were spaced approximately a nucleosome length apart, suggesting a single, regular arrangement of nucleosomes on most of the 5S DNA repeats. The observed nucleosome locations are discussed with respect to nucleotide sequences known to be important for expression of 5S RNA. Because the preferred locations appear to be reestablished in each repeating unit, despite spacer length heterogeneity, we suggest that the regular chromatin structure reflects the presence of a sequence-specific DNA-binding component on inactive 5S RNA genes.

scholarly journals Regular arrangement of nucleosomes on 5S rRNA genes in Xenopus laevis.

1983 ◽  
Vol 3 (4) ◽  
pp. 720-730 ◽  
Author(s):  
D Young ◽  
D Carroll
Keyword(s):  
Xenopus Laevis ◽  
Chromatin Structure ◽  
Micrococcal Nuclease ◽  
Rrna Genes ◽  
Dna Repeats ◽  
5S Rna ◽  
5S Dna ◽  
Regular Arrangement ◽  
5S Rna Genes ◽  
Rna Genes

The chromatin structure of the oocyte-type 5S RNA genes in Xenopus laevis was investigated. Blot hybridization analysis of DNA from micrococcal nuclease digests of erythrocyte nuclei showed that 5S DNA has the same average nucleosome repeat length, 192 +/- 4 base pairs, as two Xenopus satellite DNAs and bulk erythrocyte chromatin. The positions of nuclease-sensitive regions in the 5S DNA repeats of purified DNA and chromatin from erythrocytes were mapped by using an indirect end-labeling technique. Although most of the sites cleaved in purified DNA were also cleaved in chromatin, the patterns of intensities were strikingly different in the two cases. In 5S chromatin, three nuclease-sensitive regions were spaced approximately a nucleosome length apart, suggesting a single, regular arrangement of nucleosomes on most of the 5S DNA repeats. The observed nucleosome locations are discussed with respect to nucleotide sequences known to be important for expression of 5S RNA. Because the preferred locations appear to be reestablished in each repeating unit, despite spacer length heterogeneity, we suggest that the regular chromatin structure reflects the presence of a sequence-specific DNA-binding component on inactive 5S RNA genes.

Differential order of replication of Xenopus laevis 5S RNA genes

1986 ◽  
Vol 6 (7) ◽  
pp. 2536-2542
Author(s):  
D R Guinta ◽  
L J Korn
Keyword(s):  
Xenopus Laevis ◽  
Rna Replication ◽  
S Phase ◽  
Cold Spring Harbor ◽  
5S Rna ◽  
Expression Model ◽  
Cold Spring ◽  
5S Rna Genes ◽  
Equilibrium Centrifugation ◽  
Rna Genes

In Xenopus laevis there are two multigene families of 5S RNA genes: the oocyte-type 5S RNA genes which are expressed only in oocytes and the somatic-type 5S RNA genes which are expressed throughout development. The Xenopus 5S RNA replication-expression model of Gottesfeld and Bloomer (Cell 28:781-791, 1982) and Wormington et al. (Cold Spring Harbor Symp. Quant. Biol. 47:879-884, 1983) predicts that the somatic-type 5S RNA genes replicate earlier in the cell cycle than do the oocyte-type genes. Hence, the somatic-type 5S RNA genes have a competitive advantage in binding the transcription factor TFIIIA in somatic cells and are thereby expressed to the exclusion of the oocyte-type genes. To test the replication-expression model, we determined the order of replication of the oocyte- and somatic-type 5S RNA genes. Xenopus cells were labeled with bromodeoxyuridine, stained for DNA content, and then sorted into fractions of S phase by using a fluorescence-activated cell sorter. The newly replicated DNA containing bromodeoxyuridine was separated from the lighter, unreplicated DNA by equilibrium centrifugation and was hybridized with DNA probes specific for the oocyte- and somatic-type 5S RNA genes. In this way we found that the somatic-type 5S RNA genes replicate early in S phase, whereas the oocyte-type 5S RNA genes replicate late in S phase, demonstrating a key aspect of the replication-expression model.

scholarly journals Transcriptionally inactive oocyte-type 5S RNA genes of Xenopus laevis are complexed with TFIIIA in vitro.

1987 ◽  
Vol 7 (10) ◽  
pp. 3503-3510 ◽  
Author(s):  
L J Peck ◽  
L Millstein ◽  
P Eversole-Cire ◽  
J M Gottesfeld ◽  
A Varshavsky
Keyword(s):  
Xenopus Laevis ◽  
Differential Expression ◽  
Gene Transcription ◽  
Nuclear Extract ◽  
In Vitro Transcription ◽  
5S Rna ◽  
Differential Binding ◽  
5S Rna Genes ◽  
Rna Genes

An extract from whole oocytes of Xenopus laevis was shown to transcribe somatic-type 5S RNA genes approximately 100-fold more efficiently than oocyte-type 5S RNA genes. This preference was at least 10-fold greater than the preference seen upon microinjection of 5S RNA genes into oocyte nuclei or upon in vitro transcription in an oocyte nuclear extract. The approximately 100-fold transcriptional bias in favor of the somatic-type 5S RNA genes observed in vitro in the whole oocyte extract was similar to the transcriptional bias observed in developing Xenopus embryos. We also showed that in the whole oocyte extract, a promoter-binding protein required for 5S RNA gene transcription, TFIIIA, was bound both to the actively transcribed somatic-type 5S RNA gene and to the largely inactive oocyte-type 5S RNA genes. These findings suggest that the mechanism for the differential expression of 5S RNA genes during Xenopus development does not involve differential binding of TFIIIA to 5S RNA genes.

scholarly journals Differential order of replication of Xenopus laevis 5S RNA genes.

1986 ◽  
Vol 6 (7) ◽  
pp. 2536-2542 ◽  
Author(s):  
D R Guinta ◽  
L J Korn
Keyword(s):  
Xenopus Laevis ◽  
Rna Replication ◽  
S Phase ◽  
Cold Spring Harbor ◽  
5S Rna ◽  
Expression Model ◽  
Cold Spring ◽  
5S Rna Genes ◽  
Equilibrium Centrifugation ◽  
Rna Genes

In Xenopus laevis there are two multigene families of 5S RNA genes: the oocyte-type 5S RNA genes which are expressed only in oocytes and the somatic-type 5S RNA genes which are expressed throughout development. The Xenopus 5S RNA replication-expression model of Gottesfeld and Bloomer (Cell 28:781-791, 1982) and Wormington et al. (Cold Spring Harbor Symp. Quant. Biol. 47:879-884, 1983) predicts that the somatic-type 5S RNA genes replicate earlier in the cell cycle than do the oocyte-type genes. Hence, the somatic-type 5S RNA genes have a competitive advantage in binding the transcription factor TFIIIA in somatic cells and are thereby expressed to the exclusion of the oocyte-type genes. To test the replication-expression model, we determined the order of replication of the oocyte- and somatic-type 5S RNA genes. Xenopus cells were labeled with bromodeoxyuridine, stained for DNA content, and then sorted into fractions of S phase by using a fluorescence-activated cell sorter. The newly replicated DNA containing bromodeoxyuridine was separated from the lighter, unreplicated DNA by equilibrium centrifugation and was hybridized with DNA probes specific for the oocyte- and somatic-type 5S RNA genes. In this way we found that the somatic-type 5S RNA genes replicate early in S phase, whereas the oocyte-type 5S RNA genes replicate late in S phase, demonstrating a key aspect of the replication-expression model.

Effect of sequence differences between somatic and oocyte 5S RNA genes on transcriptional efficiency in an oocyte S150 extract

1988 ◽  
Vol 8 (11) ◽  
pp. 5056-5058
Author(s):  
W F Reynolds
Keyword(s):  
Xenopus Laevis ◽  
Differential Expression ◽  
Internal Control ◽  
Transcriptional Activity ◽  
Stable Complex ◽  
5S Rna ◽  
Internal Control Region ◽  
5S Rna Genes ◽  
Rna Genes ◽  
5S Gene

The differential expression of the Xenopus laevis somatic and oocyte 5S RNA genes is partially, but not solely, due to several base differences near the 5' boundary of the internal control region. A hybrid oocyte 5S gene with somatic-type base changes at +47, +53, +55, and +56 had intermediate transcriptional activity in oocyte S150 extracts. These base substitutions also resulted in increased affinity for a factor(s), other than TFIIIA, which forms a stable complex with the 5S gene.

The C-terminal domain of transcription factor IIIA interacts differently with different 5S RNA genes

1989 ◽  
Vol 9 (2) ◽  
pp. 499-514
Author(s):  
Y Y Xing ◽  
A Worcel
Keyword(s):  
Transcription Factor ◽  
Site Directed Mutagenesis ◽  
5S Rna ◽  
Transcription Factor Iiia ◽  
5S Dna ◽  
Type Site ◽  
Single Trace ◽  
Dna Complex ◽  
5S Rna Genes ◽  
Rna Genes

DNase I footprints and affinity measurements showed that the C-terminal arm of Xenopus transcription factor IIIA interacts differently with different Xenopus 5S DNAs, forming three distinct types of transcription factor IIIA-5S DNA complexes: a somatic type, a major-oocyte (and pseudogene) type, and a trace-oocyte type. Site-directed mutagenesis on the major-oocyte 5S gene revealed that somatic-type changes at positions 53, 55, and 56 changed the structure of the transcription factor IIIA-5S DNA complex from major-oocyte to somatic, and a single trace-oocyte change at position 56 caused the change from major-oocyte to trace-oocyte complex. We further show that the somatic-type changes are accompanied by a marked enhancement in the rate of 5S RNA transcription, and we discuss the possible biological relevance of these findings.

scholarly journals The C-terminal domain of transcription factor IIIA interacts differently with different 5S RNA genes.

1989 ◽  
Vol 9 (2) ◽  
pp. 499-514 ◽  
Author(s):  
Y Y Xing ◽  
A Worcel
Keyword(s):  
Transcription Factor ◽  
Site Directed Mutagenesis ◽  
5S Rna ◽  
Transcription Factor Iiia ◽  
5S Dna ◽  
Type Site ◽  
Single Trace ◽  
Dna Complex ◽  
5S Rna Genes ◽  
Rna Genes

DNase I footprints and affinity measurements showed that the C-terminal arm of Xenopus transcription factor IIIA interacts differently with different Xenopus 5S DNAs, forming three distinct types of transcription factor IIIA-5S DNA complexes: a somatic type, a major-oocyte (and pseudogene) type, and a trace-oocyte type. Site-directed mutagenesis on the major-oocyte 5S gene revealed that somatic-type changes at positions 53, 55, and 56 changed the structure of the transcription factor IIIA-5S DNA complex from major-oocyte to somatic, and a single trace-oocyte change at position 56 caused the change from major-oocyte to trace-oocyte complex. We further show that the somatic-type changes are accompanied by a marked enhancement in the rate of 5S RNA transcription, and we discuss the possible biological relevance of these findings.

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