Both HMG boxes in Hmo1 are essential for DNA binding <i>in vitro</i> and <i>in vivo</i>

Higashino, Ayako; Shiwa, Yuh; Yoshikawa, Hideki; Kokubo, Tetsuro; Koyamada, Koji

doi:10.1080/09168451.2014.978258

Cited by 5 publications

(13 citation statements)

References 33 publications

(55 reference statements)

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“…Based on this result, we proposed that box A, in addition to box B, is essential for DNA‐binding activity of Hmo1 (Higashino et al . ). However, another previous study carried out by others suggested that although box B is responsible for most of the DNA‐binding activity of Hmo1, box A has only a slight, if any, influence on DNA binding (Kamau et al .…”

Section: Resultsmentioning

confidence: 97%

“…For this purpose, we conducted the TSS (Transcription Start Site) shift assay, using a reporter plasmid consisting of the modified RPS5 promoter and HIS3 gene, developed in our previous study to test the DNA binding of Hmo1 in vivo (Higashino et al . ). Hmo1 binds to the intervening region between the UAS and core promoter in the modified RPS5 promoter, thereby inhibiting transcriptional initiation of ectopic mRNAs that can produce functional His3 protein.…”

Section: Resultsmentioning

confidence: 97%

“…In addition to in vivo analyses, DNA‐binding activity of Hmo1 mutant proteins was tested in vitro using an immobilized template assay, as described previously (Higashino et al . ). As expected, the DNA‐binding activity of Hmo1 mutants lacking either box A or box B was virtually abolished (Fig.…”

Section: Resultsmentioning

confidence: 97%

“…In a previous study (Higashino et al . ), we identified various mutations that significantly reduced the DNA‐binding activity of Hmo1 within aa 27–51 (e.g., T30R, A31V, S33P, V35A, F37S and N39D). Hence, it is puzzling that deletion of aa 27–51 (i.e., Δ27–51) did not significantly affect DNA binding by Hmo1.…”

Section: Resultsmentioning

confidence: 99%

“…Although we did not identify any such factors, we identified many mutations in box A, as well as in box B, that significantly decreased the DNA binding of Hmo1 (Higashino et al . ). Based on these findings, we proposed that box A is essential for DNA binding by Hmo1.…”

Section: Introductionmentioning

confidence: 97%

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Oligomerization of Hmo1 mediated by box A is essential for DNA binding in vitro and in vivo

et al. 2016

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Hmo1, a member of HMGB family proteins in Saccharomyces cerevisiae, binds to and regulates the transcription of genes encoding ribosomal RNA and ribosomal proteins. The functional motifs of Hmo1 include two HMG-like motifs, box A and box B, and a C-terminal tail. To elucidate the molecular roles of the HMG-like boxes in DNA binding in vivo, we analyzed the DNA-binding activity of various Hmo1 mutants using ChIP or reporter assays that enabled us to conveniently detect Hmo1 binding to the promoter of RPS5, a major target gene of Hmo1. Our mutational analyses revealed that box B is a bona fide DNA-binding motif, and that it also plays other important roles in cell growth. On the other hand, box A, especially its first α-helix, contributes to DNA binding of Hmo1 by inducing self-assembly of Hmo1. Intriguingly, box A mediated formation of oligomers of more than two proteins on DNA in vivo.Furthermore, duplication of the box B partially alleviates the requirement for box A. These findings suggest that the principal role of box A is to assemble multiple box B in the appropriate orientation, thereby stabilizing the binding of Hmo1 to DNA and nucleating specific chromosomal architecture on its target genes.

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Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 97%

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Oligomerization of Hmo1 mediated by box A is essential for DNA binding in vitro and in vivo

et al. 2016

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HMGB proteins involved in TOR signaling as general regulators of cell growth by controlling ribosome biogenesis

et al. 2018

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The number of ribosomes and their activity need to be highly regulated because their function is crucial for the cell. Ribosome biogenesis is necessary for cell growth and proliferation in accordance with nutrient availability and other external and intracellular signals. High-mobility group B (HMGB) proteins are conserved from yeasts to human and are decisive in cellular fate. These proteins play critical functions, from the maintenance of chromatin structure, DNA repair, or transcriptional regulation, to facilitation of ribosome biogenesis. They are also involved in cancer and other pathologies. In this review, we summarize evidence of how HMGB proteins contribute to ribosome-biogenesis control, with special emphasis on a common nexus to the target of rapamycin (TOR) pathway, a signaling cascade essential for cell growth and proliferation from yeast to human. Perspectives in this field are also discussed.

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Coupling Between Production of Ribosomal RNA and Maturation: Just at the Beginning

Azouzi

Jaafar

Dez

et al. 2021

Front. Mol. Biosci.

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Ribosomal RNA (rRNA) production represents the most active transcription in the cell. Synthesis of the large rRNA precursors (35S/47S in yeast/human) is achieved by up to hundreds of RNA polymerase I (Pol I) enzymes simultaneously transcribing a single rRNA gene. In this review, we present recent advances in understanding the coupling between rRNA production and nascent rRNA folding. Mapping of the distribution of Pol I along ribosomal DNA at nucleotide resolution, using either native elongating transcript sequencing (NET-Seq) or crosslinking and analysis of cDNAs (CRAC), revealed frequent Pol I pausing, and CRAC results revealed a direct coupling between pausing and nascent RNA folding. High density of Pol I per gene imposes topological constraints that establish a defined pattern of polymerase distribution along the gene, with a persistent spacing between transcribing enzymes. RNA folding during transcription directly acts as an anti-pausing mechanism, implying that proper folding of the nascent rRNA favors elongation in vivo. Defects in co-transcriptional folding of rRNA are likely to induce Pol I pausing. We propose that premature termination of transcription, at defined positions, can control rRNA production in vivo.

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Both HMG boxes in Hmo1 are essential for DNA binding in vitro and in vivo

Cited by 5 publications

References 33 publications

Oligomerization of Hmo1 mediated by box A is essential for DNA binding in vitro and in vivo

Oligomerization of Hmo1 mediated by box A is essential for DNA binding in vitro and in vivo

HMGB proteins involved in TOR signaling as general regulators of cell growth by controlling ribosome biogenesis

Coupling Between Production of Ribosomal RNA and Maturation: Just at the Beginning

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