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RNA pol II transcript abundance controls condensin accumulation at mitotically up-regulated and heat-shock-inducible genes in fission yeast.

Nakazawa N, Sajiki K, Xu X, Villar-Briones A, Arakawa O, Yanagida M - Genes Cells (2015)

Bottom Line: We found that condensin binds to RNA polymerase I-, II- and III-transcribed genes during both mitosis and interphase, and we focused on pol II constitutive and inducible genes.Pol II-mediated transcription was neither repressed nor activated by condensin, as levels of transcripts per se did not change when mutant condensin failed to associate with chromosomal DNA.However, massive chromosome missegregation occurred, suggesting that abundant pol II transcription may require active condensin before proper chromosome segregation.

View Article: PubMed Central - PubMed

Affiliation: G0 Cell Unit, Okinawa Institute of Science and Technology Graduate University, Onna-son, Okinawa, 904-0495, Japan.

No MeSH data available.


ChIP-seq analysis of condensin Cut14/SMC2 throughout the whole Schizosaccharomyces pombe genome in mitotic and asynchronous cells. FLAG-tagged S. pombe condensin subunit SMC2 (Cut14-FLAG) enrichment profiles along S. pombe chromosomes 1, 2, and 3 were determined using ChIP-seq. Specimens were obtained from mitotically arrested, cold-sensitive (cs) beta-tubulin mutant nda3-KM311 at the restrictive temperature 20 °C for 8 h (Mitosis, shown in orange) and asynchronously grown wild type (WT) at 20 °C (shown in blue). Ratios of precipitated DNA (IP) to input DNA are shown as the enrichment score of Cut14-FLAG-bound DNA. Representative genes are marked: cen, centromere; tR, tRNA gene; 5S, 5S rRNA gene; ncR, noncoding RNA gene; snoRNA, small nucleolar RNA; rDNA, ribosomal DNA repeats. Only 2 and 1 rDNA repeats are shown on the left and right ends of chromosome 3, respectively, because of its repetitive sequences (see text). Condensin binds to various pol II-transcribed protein-coding genes, as well as to pol I- or III-transcribed RNA genes and centromeric regions.
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fig01: ChIP-seq analysis of condensin Cut14/SMC2 throughout the whole Schizosaccharomyces pombe genome in mitotic and asynchronous cells. FLAG-tagged S. pombe condensin subunit SMC2 (Cut14-FLAG) enrichment profiles along S. pombe chromosomes 1, 2, and 3 were determined using ChIP-seq. Specimens were obtained from mitotically arrested, cold-sensitive (cs) beta-tubulin mutant nda3-KM311 at the restrictive temperature 20 °C for 8 h (Mitosis, shown in orange) and asynchronously grown wild type (WT) at 20 °C (shown in blue). Ratios of precipitated DNA (IP) to input DNA are shown as the enrichment score of Cut14-FLAG-bound DNA. Representative genes are marked: cen, centromere; tR, tRNA gene; 5S, 5S rRNA gene; ncR, noncoding RNA gene; snoRNA, small nucleolar RNA; rDNA, ribosomal DNA repeats. Only 2 and 1 rDNA repeats are shown on the left and right ends of chromosome 3, respectively, because of its repetitive sequences (see text). Condensin binds to various pol II-transcribed protein-coding genes, as well as to pol I- or III-transcribed RNA genes and centromeric regions.

Mentions: High enrichment of Cut14-FLAG was obtained for a number of chromosomal DNA sites at three chromosomes in mitosis (nda3, orange) and asynchronous (WT, blue) cells (Fig.1). Scores of condensin-enriched peaks were generally higher in nda3 samples than in WT samples. Enrichment scores for centromeres (cen1, cen2, cen3) were also high in mitosis. Most Cut14-enriched sites coincided with loci actively transcribed by pol II and pol I- or pol III-transcribed RNA genes. Some representative pol II-transcribed genes (see also Tables1 and 2, explained below) and prominent RNA genes were marked (tR, tRNA; 5S, 5S rRNA; ncR, noncoding RNA; snoRNA, small nucleolar RNA; cen1, cen2 and cen3, centromeric DNA; rDNA, ribosomal DNA). The S. pombe genome has ∽150 repeats of 10.4-kb rDNA units clustered at both ends of chromosome 3 (Schaak et al. 1982; Toda et al. 1984). Only 2 and 1 rDNA repeats were represented on the left and right ends of chromosome 3, respectively (Fig.1). These results suggest that condensin accumulates heavily at pol II-transcribed genes, in addition to previously reported condensin association sites (centromeric DNAs, pol I-transcribed rDNA repeats, pol III-transcribed tRNA and 5S rRNA genes; D'Ambrosio et al. 2008; Nakazawa et al. 2008; Iwasaki et al. 2010). We validated condensin enrichment at pol II-transcribed genes using ChIP followed by quantitative, real-time PCR (ChIP-qPCR) (Fig. S1 in Supporting Information). FLAG-tagged non-SMC subunit, Cnd1, also accumulates at pol II-transcribed genes to about the same extent as Cut14, suggesting that condensin holocomplex binds to these loci. Our ChIP-seq results for pol II-transcribed genes are in good agreement with those obtained by Drs. Takashi Sutani and Katsuhiko Shirahige (University of Tokyo, personal communication).


RNA pol II transcript abundance controls condensin accumulation at mitotically up-regulated and heat-shock-inducible genes in fission yeast.

Nakazawa N, Sajiki K, Xu X, Villar-Briones A, Arakawa O, Yanagida M - Genes Cells (2015)

ChIP-seq analysis of condensin Cut14/SMC2 throughout the whole Schizosaccharomyces pombe genome in mitotic and asynchronous cells. FLAG-tagged S. pombe condensin subunit SMC2 (Cut14-FLAG) enrichment profiles along S. pombe chromosomes 1, 2, and 3 were determined using ChIP-seq. Specimens were obtained from mitotically arrested, cold-sensitive (cs) beta-tubulin mutant nda3-KM311 at the restrictive temperature 20 °C for 8 h (Mitosis, shown in orange) and asynchronously grown wild type (WT) at 20 °C (shown in blue). Ratios of precipitated DNA (IP) to input DNA are shown as the enrichment score of Cut14-FLAG-bound DNA. Representative genes are marked: cen, centromere; tR, tRNA gene; 5S, 5S rRNA gene; ncR, noncoding RNA gene; snoRNA, small nucleolar RNA; rDNA, ribosomal DNA repeats. Only 2 and 1 rDNA repeats are shown on the left and right ends of chromosome 3, respectively, because of its repetitive sequences (see text). Condensin binds to various pol II-transcribed protein-coding genes, as well as to pol I- or III-transcribed RNA genes and centromeric regions.
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fig01: ChIP-seq analysis of condensin Cut14/SMC2 throughout the whole Schizosaccharomyces pombe genome in mitotic and asynchronous cells. FLAG-tagged S. pombe condensin subunit SMC2 (Cut14-FLAG) enrichment profiles along S. pombe chromosomes 1, 2, and 3 were determined using ChIP-seq. Specimens were obtained from mitotically arrested, cold-sensitive (cs) beta-tubulin mutant nda3-KM311 at the restrictive temperature 20 °C for 8 h (Mitosis, shown in orange) and asynchronously grown wild type (WT) at 20 °C (shown in blue). Ratios of precipitated DNA (IP) to input DNA are shown as the enrichment score of Cut14-FLAG-bound DNA. Representative genes are marked: cen, centromere; tR, tRNA gene; 5S, 5S rRNA gene; ncR, noncoding RNA gene; snoRNA, small nucleolar RNA; rDNA, ribosomal DNA repeats. Only 2 and 1 rDNA repeats are shown on the left and right ends of chromosome 3, respectively, because of its repetitive sequences (see text). Condensin binds to various pol II-transcribed protein-coding genes, as well as to pol I- or III-transcribed RNA genes and centromeric regions.
Mentions: High enrichment of Cut14-FLAG was obtained for a number of chromosomal DNA sites at three chromosomes in mitosis (nda3, orange) and asynchronous (WT, blue) cells (Fig.1). Scores of condensin-enriched peaks were generally higher in nda3 samples than in WT samples. Enrichment scores for centromeres (cen1, cen2, cen3) were also high in mitosis. Most Cut14-enriched sites coincided with loci actively transcribed by pol II and pol I- or pol III-transcribed RNA genes. Some representative pol II-transcribed genes (see also Tables1 and 2, explained below) and prominent RNA genes were marked (tR, tRNA; 5S, 5S rRNA; ncR, noncoding RNA; snoRNA, small nucleolar RNA; cen1, cen2 and cen3, centromeric DNA; rDNA, ribosomal DNA). The S. pombe genome has ∽150 repeats of 10.4-kb rDNA units clustered at both ends of chromosome 3 (Schaak et al. 1982; Toda et al. 1984). Only 2 and 1 rDNA repeats were represented on the left and right ends of chromosome 3, respectively (Fig.1). These results suggest that condensin accumulates heavily at pol II-transcribed genes, in addition to previously reported condensin association sites (centromeric DNAs, pol I-transcribed rDNA repeats, pol III-transcribed tRNA and 5S rRNA genes; D'Ambrosio et al. 2008; Nakazawa et al. 2008; Iwasaki et al. 2010). We validated condensin enrichment at pol II-transcribed genes using ChIP followed by quantitative, real-time PCR (ChIP-qPCR) (Fig. S1 in Supporting Information). FLAG-tagged non-SMC subunit, Cnd1, also accumulates at pol II-transcribed genes to about the same extent as Cut14, suggesting that condensin holocomplex binds to these loci. Our ChIP-seq results for pol II-transcribed genes are in good agreement with those obtained by Drs. Takashi Sutani and Katsuhiko Shirahige (University of Tokyo, personal communication).

Bottom Line: We found that condensin binds to RNA polymerase I-, II- and III-transcribed genes during both mitosis and interphase, and we focused on pol II constitutive and inducible genes.Pol II-mediated transcription was neither repressed nor activated by condensin, as levels of transcripts per se did not change when mutant condensin failed to associate with chromosomal DNA.However, massive chromosome missegregation occurred, suggesting that abundant pol II transcription may require active condensin before proper chromosome segregation.

View Article: PubMed Central - PubMed

Affiliation: G0 Cell Unit, Okinawa Institute of Science and Technology Graduate University, Onna-son, Okinawa, 904-0495, Japan.

No MeSH data available.