Mutations in 5S DNA and 5S RNA have different effects on the binding of Xenopus transcription factor IIIA

You, Qimin; Veldhoen, Nik; Baudin, Florence; Romaniuk, Paul J.

doi:10.1021/bi00223a028

Cited by 63 publications

(40 citation statements)

References 44 publications

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“…2B) with 3Ј boundaries at approximately positions ϩ69, ϩ80, ϩ93, and ϩ130 with respect to the transcriptional start site. TFIIIA bound to an uncomplexed X. borealis somatic 5S rRNA gene has been shown to occupy nucleotides ϩ45 to ϩ97 (32), with residues ϩ81 to ϩ91 forming the minimal requirements for TFIIIA binding (48). Thus, presumably, the fraction of mono- nucleosomes reconstituted on Xlo(Ϫ833ϩ136) which binds TFIIIA would be that with the TFIIIA binding site partially exposed (nucleosomes with 3Ј boundaries at nucleotides ϩ69 and ϩ80).…”

Section: Resultsmentioning

confidence: 99%

Nucleosome Translational Position, Not Histone Acetylation, Determines TFIIIA Binding to Nucleosomal Xenopus laevis 5S rRNA Genes

Howe

Ausió

1998

Molecular and Cellular Biology

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We sought to study the binding constraints placed on the nine-zinc-finger protein transcription factor IIIA (TFIIIA) by a histone octamer. To this end, five overlapping fragments of the Xenopus laevis oocyte and somatic 5S rRNA genes were reconstituted into nucleosomes, and it was subsequently shown that nucleosome translational positioning is a major determinant of the binding of TFIIIA to the 5S rRNA genes. Furthermore, it was found that histone acetylation cannot override the TFIIIA binding constraints imposed by unfavorable translational positions.Xenopus laevis produces two major types of 5S rRNA: the somatic type is synthesized in most cell types, whereas the oocyte type is synthesized during early oogenesis, during embryogenesis, and in certain tissue culture cell lines (14,47). Each 5S rRNA type is transcribed from a distinct multigene family, and considerable research has focused on understanding the differential expression of these genes. One hypothesis suggests that the transcription complexes which form on the oocyte genes are relatively unstable compared to those of the somatic counterparts. As a result of this, the oocyte genes are transcribed only when transcription factor IIIA (TFIIIA) levels are relatively high, as is the case during oogenesis. In somatic cells, in which TFIIIA levels are much lower, the transcription complexes dissociate from the oocyte gene, allowing the subsequent assembly of repressive nucleosome structures which preclude further factor binding (47). Conflicting with this hypothesis are results which show that, at least in vitro, the RNA polymerase III transcription complexes, once formed, have similar stabilities on both the oocyte and somatic genes (35). Thus, at this time, the reasons for the differential transcription of these two gene families in X. laevis are not fully understood.The results of 5S rRNA gene transcription studies, performed with both native and reconstituted chromatin as templates, have been conflicting. In one such study it was demonstrated that chromatin isolated from a Xenopus kidney cell line can serve as a template for transcription of the oocyte gene after the removal of histone H1 (34). In addition, it has been shown that incorporation of a somatic histone H1 variant into chromatin during embryogenesis results in specific repression of TFIIIA-activated oocyte 5S rRNA transcription (8). Furthermore, histone H1 has been shown to specifically repress transcription of oocyte genes in reconstituted chromatin while leaving the corresponding somatic genes unaffected (40). These results suggest that it is the presence of histone H1 which is responsible for the repression of oocyte transcription. This is not surprising considering that histone H1 is thought of as a repressor of transcription, although the exact mechanism of this repression is not known. In contrast to the abovementioned work, studies with chromatin reconstituted in the absence of linker histones have shown that the presence of nucleosome core particles alone is sufficient to repress oocyt...

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Section: Resultsmentioning

confidence: 99%

Nucleosome Translational Position, Not Histone Acetylation, Determines TFIIIA Binding to Nucleosomal Xenopus laevis 5S rRNA Genes

Howe

Ausió

1998

Molecular and Cellular Biology

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“…The factor makes multiple contacts over much of the nucleic acid through the nine zinc finger domains of the protein (8,9). Recognition is mediated by the secondary structure of the RNA with the major determinants located in a central domain composed of helix II-loop A-helix V-loop E (10). The other protein involved in the storage of cytoplasmic 5 S rRNA, p43, also contains nine zinc finger domains; however, it shares only 33% amino acid identity with TFIIIA (11).…”

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confidence: 99%

Analysis of the Binding of Xenopus Ribosomal Protein L5 to Oocyte 5 S rRNA

1995

Journal of Biological Chemistry

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Xenopus ribosomal protein L5 was expressed in Escherichia coli and exhibits high affinity (K d ‫؍‬ 2 nM) and specificity for oocyte 5 S rRNA. The pH dependence of the association constant for the complex reveals an ionization with a pK a value of 10.1, indicating that tyrosine and/or lysine residues are important for specific binding of L5 to the RNA. Formation of the L5⅐5 S rRNA complex is remarkably insensitive to ionic strength, providing evidence that nonelectrostatic interactions make significant contributions to binding. Together, these results suggest that one or more tyrosine residues may form critical contacts through stacking interactions with bases in the RNA. In order to locate recognition elements within 5 S rRNA, we measured binding of L5 to a collection of site-specific mutants. Mutations in the RNA that affected the interaction are confined to the hairpin structure comprised of helix III and loop C. Earlier experiments with a rhodium structural probe had shown that the two-nucleotide bulge in helix III and the intrinsic structure of loop C create sites in the major groove that are opened and accessible to stacking interactions with the metal complex. In the present studies, we detect a correlation between the intercalative binding of the rhodium complex to mutants in the hairpin and binding of L5, supporting the proposal that binding of the protein is mediated, in some part, by stacking interactions. Furthermore, the results from mutagenesis establish that, despite overlapping binding sites on 5 S rRNA, L5 and transcription factor IIIA utilize distinct structural elements for recognition.The metabolism of Xenopus 5 S ribosomal RNA during oogenesis provides the opportunity to study the interaction of several different proteins with the same nucleic acid. Moreover, these individual ribonucleoprotein complexes appear to determine the intracellular translocation of 5 S rRNA. Initially, the primary transcripts are transiently associated with the La antigen in the nucleus (1, 2). However, the synthesis of 5 S rRNA and ribosome assembly are discontinuous in the early stages of oogenesis, so that much of the RNA is stored in the cytoplasm complexed either with transcription factor IIIA (TFIIIA) 1 or with the protein p43 as part of a large multicomponent 42 S RNP complex (3, 4). Coincident with the expression of the ribosomal proteins during vitellogenesis, an increasing amount of cytoplasmic 5 S rRNA becomes associated with ribosomal protein L5 (5). This latter complex then moves to the nucleolus, where it becomes integrated into nascent ribosomes (1, 6). Mutant forms of 5 S rRNA that are unable to bind to TFIIIA or L5 are retained in the nucleus of the oocyte, indicating that the RNA can only be exported to the cytoplasm in the form of an RNP complex (2). Likewise, the ultimate return of cytoplasmic 5 S rRNA to the nucleolus depends on the formation of the complex with L5 (6, 7). It is not known how the formation of a particular RNP particle creates a signal for nucleocytoplasmic transport.The interacti...

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“…This might be due to the effects of the IE and BoxA in the affinity to the Transcription Factor IIIA (TFIIIA) compared to the BoxC. 29,30,31 It has been shown that BoxC interacts with the first 3 amino-terminal fingers of the TFIIIA, and that these are required for affinity to the 5S rDNA. 32,33 This may be the reason why BoxC is highly conserved among individuals.…”

Section: Resultsmentioning

confidence: 99%

Molecular organization of the 5S rDNA gene type II in elasmobranchs

et al. 2015

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The 5S rDNA gene is a non-coding RNA that can be found in 2 copies (type I and type II) in bony and cartilaginous fish. Previous studies have pointed out that type II gene is a paralog derived from type I. We analyzed the molecular organization of 5S rDNA type II in elasmobranchs. Although the structure of the 5S rDNA is supposed to be highly conserved, our results show that the secondary structure in this group possesses some variability and is different than the consensus secondary structure. One of these differences in Selachii is an internal loop at nucleotides 7 and 112. These mutations observed in the transcribed region suggest an independent origin of the gene among Batoids and Selachii. All promoters were highly conserved with the exception of BoxA, possibly due to its affinity to polymerase III. This latter enzyme recognizes a dT 4 sequence as stop signal, however in Rajiformes this signal was doubled in length to dT 8. This could be an adaptation toward a higher efficiency in the termination process. Our results suggest that there is no TATA box in elasmobranchs in the NTS region. We also provide some evidence suggesting that the complexity of the microsatellites present in the NTS region play an important role in the 5S rRNA gene since it is significantly correlated with the length of the NTS.

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Mutations in 5S DNA and 5S RNA have different effects on the binding of Xenopus transcription factor IIIA

Cited by 63 publications

References 44 publications

Nucleosome Translational Position, Not Histone Acetylation, Determines TFIIIA Binding to Nucleosomal Xenopus laevis 5S rRNA Genes

Nucleosome Translational Position, Not Histone Acetylation, Determines TFIIIA Binding to Nucleosomal Xenopus laevis 5S rRNA Genes

Analysis of the Binding of Xenopus Ribosomal Protein L5 to Oocyte 5 S rRNA

Molecular organization of the 5S rDNA gene type II in elasmobranchs

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