Epigenetic Chromatin Regulation

  • Small RNA-mediated transcriptional gene silencing is a fundamental process that has been observed in many different eukaryotes including fungi, plants, flies, worms and mammals. One of its main tasks is to neutralize the activity of transposable elements (TEs), which otherwise destabilize our genome and potentially cause diseases. Small RNAs use their base complementarities to TEs to specifically silence them. However, how cells ensure TE-silencing without disturbing expressions of normal genes is not fully understood.

    The ciliated protozoan Tetrahymena identifies TE-derived sequences by a germline-some genome comparison mechanism using small RNAs during programmed DNA elimination, which provides fascinating examples of epigenetic genome regulations and important insights into the interaction between TEs and host genomes. Because programmed DNA elimination can be synchronously induced in laboratory in a large scale, it serves as a useful laboratory model for genetically and biochemically investigating small RNA-mediated chromatin regulation.

    Using this tiny-hairy eukaryotic model, we aim to understand: how cells accumulate small RNAs specifically from TE-related sequences; how cells use those small RNAs to identify TE-related sequences; and how a small RNA pathway establishes silent chromatin environment (heterochromatin) on TE-related sequences.

    Figure 1
    Tetrahymena thermophila. Tetrahymena is a unicellular eukaryote. Tetrahymena has many cilia on its cell surface (green = anti-alpha tubulin staining). Tetrahymena has two different nuclei (stained purple), the smaller germline micronucleus (MIC) and the larger somatic macronucleus (MAC).

    Members

    Kazufumi Mochizuki
    Mochizuki Kazufumi
    Julie Saksouk
    Saksouk Julie
    Masatoshi Mutazono
    Mutazono Masatoshi
    Tomoko Noto
    Noto Tomoko
    Eliot Geraud
    Geraud Eliot
    Salman Shehzada
    Shehzada Salman

    Publications

    Diversification of small RNA amplification mechanisms for targeting transposon-related sequences in ciliates.

    Mutazono M, Noto T, Mochizuki K.

    2019 - Proc Natl Acad Sci USA

    Request for full article31262823

    Small RNA-Mediated trans-Nuclear and trans-Element Communications in Tetrahymena DNA Elimination.

    Noto T, Mochizuki K

    2018 - Curr Biol

    Request for full article29887308

    Phosphorylation of an HP1-like Protein Regulates Heterochromatin Body Assembly for DNA Elimination.

    Kataoka K, Mochizuki K

    2015 - Dev Cell, 35(6):775-88

    Request for full article26688337

    Small-RNA-Mediated Genome-wide trans-Recognition Network in Tetrahymena DNA Elimination.

    Noto T, Kataoka K, Suhren JH, Hayashi A, Woolcock KJ, Gorovsky MA, Mochizuki K

    2015 - Mol Cell, 59(2):229-42

    Request for full article26095658
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    Publications of the team

  • Background: Small RNA-directed DNA elimination of Tetrahymena

    The ciliated protozoan Tetrahymena possesses a somatic macronucleus (MAC) and a germline micronucleus (MIC) in each cell. MAC is polyploid and transcriptionally active, whereas MIC is diploid and transcriptionally inert during vegetative growth. In the sexual reproduction, MIC gives rise to a new MAC and a new MIC, and the parental MAC is destroyed. During the development of the new MAC, ~12,000 internal eliminated sequences (IESs) are removed (DNA elimination) and the remaining MAC-destined sequences are re-ligated. Most IESs are moderately repeated in the MIC and many of them are related to transposable elements. Small RNA-directed heterochromatin formation is involved in the DNA elimination process. Small (~29 nt) RNAs, called Early-scnRNAs, are produced by the Dicer protein Dcl1p, and associate with the Argonaute protein Twi1p. Early-scnRNAs induce accumulation of heterochromatin components, including histone H3 methylated on lysine 9 (H3K9me) and on lysine 27 (H3K27me), and the chromodomain protein Pdd1p, specifically on IES sequences. Heterochromatin eventually recruits the endonuclease Tpb2p, which catalyzes DNA elimination.

    Figure 2
    Small RNA-directed heterochromatin formation induces DNA elimination. Non-coding (nc) RNAs derived from the MIC genome, including transposons, are processed to Early-scnRNAs by Dcl1p (a). Early-scnRNA forms a complex with the Argonaute protein Twi1p (b). Ema1p facilitates interaction between the complex and nascent MAC ncRNA (c). This interaction recruits Ezl1p (d), which catalyzes methylations of histone H3 at lys9 and lys27 (e). Pdd1p and Pdd3p bind to the methylated histone H3 and establish heterochromatin structure (f). Tpb2p mediates the final DNA excision process (g).

    Epigenetic regulation of DNA elimination by small RNA-mediated germline-soma genome comparison

    The fact that IESs do not share any common sequence motifs raises the following question: how is Tetrahymena able to identify IESs to induce DNA elimination? Tetrahymena solves this problem by germline-soma genome comparison. In a single cell, Tetrahymena has a germline MIC, which contains complete genome including IESs, and a somatic MAC in which IESs are removed during the last sexual reproduction. Thus, the cell is able to identify IESs as sequences existing in the MIC but not in the MAC. Tetrahymena utilizes Early-scnRNAs for this transnuclear genome comparison. This system sweeps away not only the existing TEs, but also any newly invaded TEs from the transcriptionally active MAC. We previously reported that only Early-scnRNAs complementary to the parental MAC genome are degraded during sexual reproduction, and this selective turnover mediates the germline-soma genome comparison. We are currently trying to understand the molecular mechanism behind the selective turnover of Early-scnRNAs.

    Small RNA-mediated genome-wide trans-recognition network for DNA elimination

    In addition to the germline-soma transnuclear genome comparison mechanism explained above, we recently reported that an additional mechanism of small RNA-mediated identification of TE-related sequences. We showed that only ~60% of IESs, called Type-A IESs, produces Early-scnRNAs. Early-scnRNAs recognize not only Type-A IESs from which they are derived but also other ~40% of IESs, called Type-B IESs, in trans. This trans-recognition triggers the expression of yet another ~29-nt RNAs, named Late-scnRNAs, that further identify other IESs. Therefore, IESs in Tetrahymena are robustly targeted for DNA elimination by a genome-wide trans-recognition network accompanied by a chain reaction of small RNA production. We found that the biogenesis of Late-scnRNAs require heterochromatin components. We are going to understand how heterochromatin, which generally inhibit transcription, induces production of Late-scnRNAs.

    Figure 3
    Germline-soma transnuclear genome comparison and trans-recognition network in DNA elimination. Early-scnRNAs are expressed from Type-A IESs (magenta boxes) and their flanking sequences in the MIC (i), and the latter are degraded in the parental MAC (ii). In the new MAC, Early-scnRNAs recognize the IESs from which they are derived (iii) as well as other Type-A and Type-B (sky-blue boxes) IESs in trans (iv) through common repeats to trigger Late-scnRNA production (v). In an IES, regions producing Late-scnRNAs spread in cis (vi) in heterochromatin component (Pdd1p, H3K9me)-dependent manner.
  • Education

    2000 - Ph. D, The Graduate University for Advanced Studies, School of Life Science, Department of Genetics, Japan
    1997 - Master of Engineering, Tokyo Institute of Technology, School of Bioscience and Biotechnology, Japan
    1995 - Bachelor of Engineering, Tokyo Institute of Technology, Department of Bioscience and Biotechnology, Japan

    Postdoctoral Training and Employment History

    2016-present: Group Leader (CNRS, DR2), Institute of Human Genetics (IGH), Montpellier, France
    2006-2016: Group Leader, Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna, Austria
    2001-2005: Postdoctoral fellow, University of Rochester, Laboratory of Prof. Martin A. Gorovsky, Rochester, NY, USA
    2000-2001: Postdoctoral fellow, National Institute of Genetics, Laboratory of Prof. Toshitaka Fujisawa, Mishima, Japan