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The yeast prefoldin-like URI-orthologue Bud27 associates with the RSC nucleosome remodeler and modulates transcription.

Mirón-García MC, Garrido-Godino AI, Martínez-Fernández V, Fernández-Pevida A, Cuevas-Bermúdez A, Martín-Expósito M, Chávez S, de la Cruz J, Navarro F - Nucleic Acids Res. (2014)

Bottom Line: In this work, we present evidence of Bud27 modulating RNA pol II transcription elongation.We also reveal that Bud27 associates in vivo with the Sth1 component of the chromatin remodeling complex RSC and mediates its association with RNA pol II.Our data suggest that Bud27, in addition of contributing to Rpb5 folding within the RNA polymerases, also participates in the correct assembly of other chromatin-associated protein complexes, such as RSC, thereby modulating their activity.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biología Experimental, Facultad de Ciencias Experimentales, Universidad de Jaén, Paraje de las Lagunillas, s/n, 23071, Jaén, Spain.

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BUD27 and RPB5 genetically interact. (A) List of genetic interactions between rpb5 and bud27Δ mutants based on their growth in YPD plates. (B) Left panel, schematic view of Rpb5 (purple) and the cleft domain (orange) of the RNA pol II of S. cerevisiae on the structure of the complex, and the position of Rpb5 residues whose point mutations affect (blue) or do not affect growth (cyan). Right panel, zoom view of the section indicated in the left panel.
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Figure 1: BUD27 and RPB5 genetically interact. (A) List of genetic interactions between rpb5 and bud27Δ mutants based on their growth in YPD plates. (B) Left panel, schematic view of Rpb5 (purple) and the cleft domain (orange) of the RNA pol II of S. cerevisiae on the structure of the complex, and the position of Rpb5 residues whose point mutations affect (blue) or do not affect growth (cyan). Right panel, zoom view of the section indicated in the left panel.

Mentions: Bud27/URI physically binds different nuclear proteins involved in transcription, among them Rpb5, a shared subunit of the three nuclear RNA polymerases (3,5,7,11,12,15). However, the genetic interaction between BUD27 and RPB5 in S. cerevisiae has never been examined. We tested for conditional synthetic interactions between 25 previously generated rpb5 mutations (25) and the bud27Δ allele. As shown in Figure 1A, 15 out of the 25 double mutants were lethal or aggravated their growth with respect to the single mutants. Notably, as shown in Figure 1B, all the rpb5 point mutations affecting growth of the bud27Δ strain lie at the C-terminal globe of Rpb5. This motif, which is conserved from archaea to higher eukaryotes, is involved in the interaction of Rpb5 with Rpb1, in a region directly associated with the RNA pol II cleft domain (α45–47 of Rpb1). In addition to some yeast-human chimeric RPB5 alleles, the other rpb5 mutations (Δ10, Δ12 and Δ14) exhibiting genetic interaction with bud27Δ corresponded to a different highly conserved motif (positions 11–30), belonging to the long hydrophilic helix Rpb5-α1 that occupies the ‘lower’ far-end of the DNA cleft (25,26). These results are consistent with our previous data showing that RPB5 overexpression suppresses the thermosensitivity and rapamycin-sensitivity phenotypes of the bud27Δ strain (3). It bears noting that these genetic interactions reflect the participation of both proteins in common biological process and seems not to be the consequence of a loss of physical contact between Rpb5 and Bud27, as we demonstrated by two-hybrid analyses (Supplementary Figure S1).


The yeast prefoldin-like URI-orthologue Bud27 associates with the RSC nucleosome remodeler and modulates transcription.

Mirón-García MC, Garrido-Godino AI, Martínez-Fernández V, Fernández-Pevida A, Cuevas-Bermúdez A, Martín-Expósito M, Chávez S, de la Cruz J, Navarro F - Nucleic Acids Res. (2014)

BUD27 and RPB5 genetically interact. (A) List of genetic interactions between rpb5 and bud27Δ mutants based on their growth in YPD plates. (B) Left panel, schematic view of Rpb5 (purple) and the cleft domain (orange) of the RNA pol II of S. cerevisiae on the structure of the complex, and the position of Rpb5 residues whose point mutations affect (blue) or do not affect growth (cyan). Right panel, zoom view of the section indicated in the left panel.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4150788&req=5

Figure 1: BUD27 and RPB5 genetically interact. (A) List of genetic interactions between rpb5 and bud27Δ mutants based on their growth in YPD plates. (B) Left panel, schematic view of Rpb5 (purple) and the cleft domain (orange) of the RNA pol II of S. cerevisiae on the structure of the complex, and the position of Rpb5 residues whose point mutations affect (blue) or do not affect growth (cyan). Right panel, zoom view of the section indicated in the left panel.
Mentions: Bud27/URI physically binds different nuclear proteins involved in transcription, among them Rpb5, a shared subunit of the three nuclear RNA polymerases (3,5,7,11,12,15). However, the genetic interaction between BUD27 and RPB5 in S. cerevisiae has never been examined. We tested for conditional synthetic interactions between 25 previously generated rpb5 mutations (25) and the bud27Δ allele. As shown in Figure 1A, 15 out of the 25 double mutants were lethal or aggravated their growth with respect to the single mutants. Notably, as shown in Figure 1B, all the rpb5 point mutations affecting growth of the bud27Δ strain lie at the C-terminal globe of Rpb5. This motif, which is conserved from archaea to higher eukaryotes, is involved in the interaction of Rpb5 with Rpb1, in a region directly associated with the RNA pol II cleft domain (α45–47 of Rpb1). In addition to some yeast-human chimeric RPB5 alleles, the other rpb5 mutations (Δ10, Δ12 and Δ14) exhibiting genetic interaction with bud27Δ corresponded to a different highly conserved motif (positions 11–30), belonging to the long hydrophilic helix Rpb5-α1 that occupies the ‘lower’ far-end of the DNA cleft (25,26). These results are consistent with our previous data showing that RPB5 overexpression suppresses the thermosensitivity and rapamycin-sensitivity phenotypes of the bud27Δ strain (3). It bears noting that these genetic interactions reflect the participation of both proteins in common biological process and seems not to be the consequence of a loss of physical contact between Rpb5 and Bud27, as we demonstrated by two-hybrid analyses (Supplementary Figure S1).

Bottom Line: In this work, we present evidence of Bud27 modulating RNA pol II transcription elongation.We also reveal that Bud27 associates in vivo with the Sth1 component of the chromatin remodeling complex RSC and mediates its association with RNA pol II.Our data suggest that Bud27, in addition of contributing to Rpb5 folding within the RNA polymerases, also participates in the correct assembly of other chromatin-associated protein complexes, such as RSC, thereby modulating their activity.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biología Experimental, Facultad de Ciencias Experimentales, Universidad de Jaén, Paraje de las Lagunillas, s/n, 23071, Jaén, Spain.

Show MeSH
Related in: MedlinePlus