The yeast prefoldin-like URI-orthologue Bud27 associates with the RSC nucleosome remodeler and modulates transcription.
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.
Affiliation: Departamento de Biología Experimental, Facultad de Ciencias Experimentales, Universidad de Jaén, Paraje de las Lagunillas, s/n, 23071, Jaén, Spain.Show MeSH
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Mentions: It has been reported that Rpb5 physically interacts with the Rsc4 subunit of RSC complex, an essential multisubunit chromatin remodeling complex that participates in transcription elongation (32,33). Moreover, certain thermosensitive rpb5 alleles are lethal in combination with a specific rsc4 mutation (rsc4-Δ4), supporting the physiological significance of the Rpb5-Rsc4 interaction (18). Genetic interactions between BUD27 and RSC1 and RSC8, encoding other components of RSC, has been reported (34). In addition, Bud27 has been shown to genetically or physically associate with other elements of the yeast chromatin machinery, such as Paf1, SAGA, histone deacetylases or SWR1, among others (34–37). Based on these findings, we investigated a possible physical interaction between Bud27 and the RSC complex. We first analysed the association between Bud27 and RSC complex by performing protein co-immunoprecipitation. To do so, we introduced a functional Myc-tagged version of the Sth1 subunit of the RSC complex (18), in a bud27Δ strain. We also expressed in this strain a functional Bud27-GFP protein from a plasmid (3). The bud27Δ strain transformed with a vector expressing Bud27-GFP was used as untagged control. As shown in Figure 5A, when Sth1-Myc was immunoprecipitated in the strain expressing Bud27-GFP (Green Fluorescent Protein), an anti-GFP reacting band was detected (line 2). In clear contrast, no such band was observed when the immunoprecipitation was performed in the control strain expressing Bud27-GFP in an untagged Sht1 background (No tag). In addition, no anti-Sth1-Myc or anti-Bud27-GFP-reacting material was adsorbed non-specifically onto the beads (Nc). These observations indicate that Bud27 specifically interacts with the Sth1 subunit of RSC complex, and probably therefore with RSC.
Affiliation: Departamento de Biología Experimental, Facultad de Ciencias Experimentales, Universidad de Jaén, Paraje de las Lagunillas, s/n, 23071, Jaén, Spain.