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Gene regulation by the lysine demethylase KDM4A in Drosophila
Stockholm University, Faculty of Science, The Wenner-Gren Institute.
Stockholm University, Faculty of Science, The Wenner-Gren Institute.
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2013 (English)In: Developmental Biology, ISSN 0012-1606, E-ISSN 1095-564X, Vol. 373, no 2, 453-463 p.Article in journal (Refereed) Published
Abstract [en]

Lysine methylation of histones is associated with both transcriptionally active chromatin and with silent chromatin, depending on what residue is modified. Histone methyltransferases and demethylases ensure that histone methylations are dynamic and can vary depending on cell cycle- or developmental stage. KDM4A demethylates H3K36me3, a modification enriched in the 3' end of active genes. The genomic targets and the role of KDM4 proteins in development remain largely unknown. We therefore generated KDM4A mutant Drosophila, and identified 99 mis-regulated genes in first instar larvae. Around half of these genes were down-regulated and the other half up-regulated in dKDM4A mutants. Although heterochromatin protein 1a (HP1a) can stimulate dKDM4A demethylase activity in vitro, we find that they antagonize each other in control of dKDM4A-regulated genes. Appropriate expression levels for some dKDM4A-regulated genes rely on the demethylase activity of dKDM4A, whereas others do not. Surprisingly, although highly expressed, many demethylase-dependent and independent genes are devoid of H3K36me3 in wild-type as well as in dKDM4A mutant larvae, suggesting that some of the most strongly affected genes in dKDM4A mutant animals are not regulated by H3K36 methylation. By contrast, dKDM4A over-expression results in a global decrease in H3K36me3 levels and male lethality, which might be caused by impaired dosage compensation. Our results show that a modest increase in global H3K36me3 levels is compatible with viability, fertility, and the expression of most genes, whereas decreased H3K36me3 levels are detrimental in males.

Place, publisher, year, edition, pages
2013. Vol. 373, no 2, 453-463 p.
Keyword [en]
Chromatin, Histone methylation, Gene regulation, Drosophila
National Category
Developmental Biology
Research subject
Developmental Biology
Identifiers
URN: urn:nbn:se:su:diva-85852DOI: 10.1016/j.ydbio.2012.11.011ISI: 000313381200020PubMedID: 23195220OAI: oai:DiVA.org:su-85852DiVA: diva2:585159
Available from: 2013-01-09 Created: 2013-01-09 Last updated: 2017-12-06Bibliographically approved
In thesis
1. Regulators of chromatin and transcription in Drosophila
Open this publication in new window or tab >>Regulators of chromatin and transcription in Drosophila
2013 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Development of multicellular organisms is achieved by organized temporal and spatial patterns of gene expression leading to cell differentiation. Chromatin regulators control how the DNA is utilized by altering access of proteins to DNA and thereby function as co-factors in transcription. Gene regulation also involves co-factors interacting with transcription factors at regulatory sequences of DNA. In this thesis, we have studied the in vivo role of three co-factors, CBP, dKDM4A and Brakeless, in regulating chromatin and transcription using Drosophila melanogaster. The CREB binding protein (CBP) belongs to histone acetyl transferases (HATs) and facilitates gene activation by many transcription factors. Our work has demonstrated that CBP occupies the genome preferentially together with Rel and Smad proteins controlling dorsal-ventral patterning in the Drosophila embryo. CBP occupancy generally correlates with gene expression but also occurs at silent genes without resulting in histone acetylation. KDM4A belongs to a family of JmjC domain proteins and demethylates H3K36me3, a histone modification enriched in the 3’end of active genes. We generated dKDM4A mutants with a global elevation of H3K36me3 levels and identify mis-regulated genes in first instar larvae. The data indicate that dKDM4A regulates some genes by mechanisms that do not involve H3K36 methylation. Further, over-expression of dKDM4A result in male lethality and globally reduced H3K36me3 levels, indicating impaired dosage compensation of the X-chromosome. Brakeless is a conserved co-factor participating in several important processes during development. We generated mutant brakeless embryos and identify direct genomic targets of Brakeless. To our surprise, Brakeless behaves as a direct activator for some genes but repressor in other cases. We also identify an interaction of Brakeless with the Mediator subunit Med19. In summary, these studies reveal unexpected roles for co-regulators in Drosophila development. The HAT CBP can bind silent genes without leading to histone acetylation. Brakeless has the ability to function both as a direct activator and repressor of transcription.

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 2013. 65 p.
Keyword
Transcription, chromatin, co-regulators, CBP, lysine demethylase, KDM4A, Brakeless, Drosophila
National Category
Developmental Biology
Research subject
Developmental Biology
Identifiers
urn:nbn:se:su:diva-87378 (URN)978-91-7447-646-0 (ISBN)
Public defence
2013-03-08, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 13:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 3: Manuscript.

Available from: 2013-02-14 Created: 2013-02-04 Last updated: 2013-03-20Bibliographically approved
2. Gene regulation during development by chromatin and the Super Elongation Complex
Open this publication in new window or tab >>Gene regulation during development by chromatin and the Super Elongation Complex
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Developmental processes are carefully controlled at the level of transcription to ensure that the fertilized egg develops into an adult organism. The mechanisms that controls transcription of protein-coding genes ultimately ensure that the Pol II machine synthesizes mRNA from the correct set of genes in every cell type. Transcriptional control involves Pol II recruitment as well as transcriptional elongation. Recent genome-wide studies shows that recruitment of Pol II is often followed by an intermediate step where Pol II is halted in a promoter-proximal paused configuration. The release of Pol II from promoter-proximal pausing is thus an additional and commonly occurring mechanism in metazoan gene regulation. The serine kinase P-TEFb is part of the Super Elongation Complex that regulates the release of paused Pol II into productive elongation. However, little is known about the role of P-TEFb mediated gene expression in development. We have investigated the function of P-TEFb in early Drosophila embryogenesis and find that P-TEFb and other Super Elongation Complex subunits are critical for activation of the most early expressed genes. We demonstrate an unexpected function for Super Elongation Complex in activation of genes with non-paused Pol II. Furthermore, the Super Elongation Complex shares phenotypes with subunits of the Mediator complex to control the activation of essential developmental genes. This raises the possibility that the Super Elongation Complex has an unappreciated role in the recruitment of Pol II to promoters. The unique chromatin landscape of each cell type is comprised of post-translational chromatin modifications such as histone methylations and acetylations. To study the function of histone modifications during development, we depleted the histone demethylase KDM4A in Drosophila to evaluate the role of KDM4A and histone H3 lysine 36 trimethylation (H3K36me3) in gene regulation. We find that KDM4A has a male-specific function and regulates gene expression both by catalytic-dependent and independent mechanisms. Furthermore, we used histone replacement to investigate the direct role of H3K14 acetylation in a multicellular organism. We show that H3K14 acetylation is essential for development, but is not cell lethal, suggesting that H3K14 acetylation has a critical role in developmental gene regulation. This work expands our knowledge of the mechanisms that precisely controls gene regulation and transcription, and in addition highlights the complexity of metazoan development.

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 2014. 58 p.
Keyword
Drosophila development, gene regulation, transcription, chromatin, P-TEFb, Super Elongation Complex, KDM4A, H3K36me3, H3K14ac, histone replacement
National Category
Developmental Biology
Research subject
Developmental Biology
Identifiers
urn:nbn:se:su:diva-103066 (URN)978-91-7447-932-4 (ISBN)
Public defence
2014-06-05, sal E306, Arrheniuslaboratorierna, Svante Arrhenius väg 20 C, Stockholm, 13:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 1: Manuscript. Paper 3: Manuscript.

Available from: 2014-05-14 Created: 2014-04-30 Last updated: 2014-05-07Bibliographically approved

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