Ensinck, Imke Maria Franciscus;
(2023)
Dissecting the N6-methyladenosine writer complex in budding yeast.
Doctoral thesis (Ph.D), UCL (University College London).
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Abstract
Precise regulation of gene expression is vital for all biological processes, and takes places at multiple levels including transcription, RNA metabolism, and posttranslational modification. In addition, mRNA molecules can be modified which represents an additional layer of gene expression regulation. The most abundant mRNA modification is N6-methyladenosine (m6A). This modification affects various biochemical reactions including mRNA decay and translation. Moreover, m6A is implicated in a wide range of physiological processes such as differentiation and gametogenesis, and dysregulation of the m6A modification has been associated with multiple types of cancer and other diseases. The m6A mark is deposited on mRNA transcripts by methyltransferase complexes known as m6A writers. While the biochemical reaction to form m6A-modified RNAs is well characterized in vitro, m6A deposition in vivo remains poorly understood. Here, I used the budding yeast Saccharomyces cerevisiae to dissect how m6A deposition is regulated. In budding yeast, m6A is only deposited during early meiosis. Using a multifaceted approach, including proteomics, yeast genetics, genomics, and a new method to rapidly determine relative m6A levels across mRNA samples (m6A-ELISA), I identified and characterized the yeast m6A writer complex. I found that the yeast m6A writer consists of six proteins (Ime4, Mum2, Slz1, Kar4, YGL036W, Dyn2), of which 5 are essential for m6A deposition (Ime4, Mum2, Slz1, Kar4, YGL036W), and of which three were previously known (Ime4, Mum2, Slz1). Sequence and structure prediction data reveal that the m6A writer is highly similar between yeast, plants, and mammals. Apart from m6A deposition, several of the writer components have additional cellular functions: Kar4 has a separate function in mating, and a subcomplex (Ime4, Mum2, YGL036W, and Kar4) promotes meiosis in an m6A-independent manner. My work forms a starting point for understanding the molecular basis for m6A deposition as well as new molecular functions of m6A writer complexes.
| Type: | Thesis (Doctoral) |
|---|---|
| Qualification: | Ph.D |
| Title: | Dissecting the N6-methyladenosine writer complex in budding yeast |
| Open access status: | An open access version is available from UCL Discovery |
| Language: | English |
| Additional information: | Copyright © The Author 2023. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request. |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences |
| URI: | https://discovery-pp.ucl.ac.uk/id/eprint/10184466 |
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