Studying stem cells for the future of regenerative medicine
transcription factors – SOX2 and OCT4 – are thought to play a major role in this respect. “We believe that these are the first players that act at the Mitosis-G1 transition to activate genes in pluripotent stem cells. The first thing they need to do is to de-condense DNA that is condensed during mitosis, but in the right places,” says Professor Suter. “This enables the re-activation of the sequences controlling the activity of genes which are important in stem cell self-renewal.”
OCT4 and SOX2 A variety of techniques are being applied in research, including a method called ChIPSeq, which enables researchers to map where OCT4 and SOX2 bind to the genome. This represents an important step towards the reactivation of gene transcription. “Our current thinking is that after mitosis, SOX2 and OCT4 bind first to genes involved in selfrenewal. Then they also need other factors
between different parts of these long strings. These contacts are very important for the regulation of gene expression, so they have to be re-established. Once they have been reestablished, then other factors can also come in.” While the project is mainly focused on ESCs at this stage, Professor Suter hopes that this research will hold broader relevance to different types of stem cells. “Once we’ve identified all the players, and the right sequence of events, we hope that this is going to be applicable to different kinds of stem cells,” he continues. This research represents an important step towards the long-term goal of producing cells directly for regenerative medicine, which has generated a lot of interest as a means of treating conditions like Parkinson’s disease or heart infarction. However, while this is an exciting prospect, Professor Suter says that it is essential to build a deeper understanding of
We develop tools to quantitatively study gene expression in living cells. So we can follow – in real time – the expression of messenger RNA or proteins.
One of the most crucial attributes of stem cells is their capacity to self-renew, in other words their ability to maintain the same identity even after undergoing cell division. Professor David Suter tells us about his work in investigating the capacity of embryonic stem cells to self-renew, which could in future help open up new possibilities in regenerative medicine. The process of cell division has a considerable influence on gene expression, which plays a major role in determining the identity of a cell, topics central to Professor David Suter’s research agenda. As the head of a biology laboratory at EPFL in Lausanne, Professor Suter is particularly interested in the capacity of embryonic stem cells (ESCs) to self-renew. “This means that they can divide and maintain the same identity, the same phenotype. We use ESCs as a model system to investigate this property of selfrenewal, which is very important for all types of stem cells,” he outlines. Both ESCs and induced pluripotent stem cells (IPSCs) can be transformed into virtually any cell type of the body, so offer great potential in terms of regenerative medicine, a major motivating factor behind Professor Suter’s research. “In principle, these cells can renew themselves to replace those that are lost in some diseases,” he explains. Self-renewal A further important motivation behind this research is a fundamental interest in the
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underlying mechanisms behind self-renewal and the nature of the process. Professor Suter is particularly interested in what occurs in cells during and after mitosis, the point in the cell cycle where chromosomes separate. “During mitosis chromosomes are condensed and genes are essentially shut off. When cells exit mitosis, they need to reactivate all the correct genes to maintain their identity. If they don’t reactivate the right genes, then the cell identity will change,” he explains. When a cell exits mitosis, a first wave of gene expression to turn on the genes specific to that cell identity occurs. “The whole molecular architecture that allows genes to be directly expressed is basically destroyed during mitosis. This architecture effectively needs to be reconstructed when a cell exits mitosis,” says Professor Suter. There are many open questions around this process, now Professor Suter and his colleagues aim to build a deeper understanding of the underlying mechanisms. This work involves analysing both single cells and larger populations, although one of the main strengths of Professor Suter’s lab is in
looking at single cells, mainly using live-cell imaging. “We develop tools to quantitatively study gene expression in living cells. So we can follow – in real time – the expression of messenger RNA or proteins. We can quantify these to understand how this actually affects stem cells and to track their phenotypic state in real time,” he explains. Researchers in the group are studying ESCs from mice. “These cells are taken from the embryo at the blastocyst stage,” says Professor Suter. “Once we have taken these cells, we maintain them in a petri dish, and we can then make them divide as much as desired.” These cells can be separated in different phases of the cell cycle through the use of flow cytometry, then different methods can be applied to look at them in greater depth. Researchers are investigating which are the main players in re-activating genes following the exit from mitosis to the G1 phase of the cell cycle, which is known as the Mitosis-G1 transition. “Which are the main molecules involved in re-activating genes in early G1? How are these molecules coordinated ?” outlines Professor Suter. Two pluripotency
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to help them to reactivate transcription. So we think there’s a whole series of molecules that act one after the other in a coordinated manner to allow stem cell self-renewal,” says Professor Suter. Researchers are also using a technique called ATAC-seq to identify where decondensation happens in the whole genome. “We use this method to determine how OCT4 binds to DNA and opens chromatin during mitotic exit, and to investigate any other changes. We’ve found that there are many places essential for stem cell self-renewal that do not recover their accessibility after mitosis in the absence of OCT4,” continues Professor Suter. Furthermore; “DNA in the nucleus is not randomly organised, there are contacts
how cells self-renew. “We want to understand how cells control their identity through cell division. As long as we don’t understand this, it’s going to be difficult to control,” he stresses. There is still much to learn in this respect, and the events around the Mitosis-G1 transition will remain an important part of Professor Suter’s research agenda over the coming years. Changes in chromatin accessibility as a function of OCT4 level decrease over time using inducible OCT4 degradation, at a genomic location where OCT4 binds (OCT4 ChIP).
Changes in chromatin accessibility of different gene regulatory sequences upon loss of OCT4 at the Mitosis-G1 transition (red lines). Some regions of the genome are strongly dependent on the presence of OCT4 upon mitotic exit (Cluster 1, left), while other regions are independent of OCT4 (Cluster 4, right). Regions dependent on OCT4 are enriched in elements regulating stem cell selfrenewal. EG1: early G1 phase; LG1: late G1 phase; S: S phase; SG2: end of S phase and G2 phase.
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Mechanisms of cell-cycle dependent cell fate regulation by OCT4 Project Objectives
The objectives of the laboratory are; i) To develop and apply quantitative, timeresolved approaches to gene expression and gene regulation, with a strong focus on single cell live microscopy; ii) To apply these methods to address central questions in gene expression regulation and stem cell biology.
Project Funding
Funded by the Swiss National Science Foundation (SNSF)
Contact Details
Project Coordinator, Professor David Suter SNSF and Tenure Track Assistant Professor Institute of Bioengineering Ecole Polytechnique Federale de Lausanne (EPFL) Switzerland T: +41 21 693 96 31 E: david.suter@epfl.ch W: http://p3.snf.ch/project-179068 Phillips NE*, Mandic A*, Omidi S, Naef F†, Suter DM†. Memory and relatedness of transcriptional activity in mammalian cell lineages. *,†Equal contribution. Nature Communications 2019 March 14. Raccaud M, Alber AB, Friman ET, Agarwal H, Deluz C, Kuhn T, Gebhardt JCM, Suter DM. Mitotic chromosome binding predicts transcription factor properties in interphase. Nature Communications 2019 Jan 30. Alber AB*, Paquet ER*, Biserni M, Naef F, Suter DM. Single Live Cell Monitoring of Protein Turnover Reveals Intercellular Variability and Cell-Cycle Dependence of Degradation Rates. Molecular Cell 2018 Aug 23. Deluz C*, Friman ET*, Strebinger D*, Benke A, Raccaud M, Callegari A, Leleu M, Manley S, Suter DM. A role for mitotic bookmarking of SOX2 in pluripotency and differentiation. Genes & Development 2016 Dec 5. Friman ET, Deluz C, Meireles-Filho A, Govindan A, Gardeux V, Deplancke B, Suter DM. Dynamic regulation of chromatin accessibility by pluripotency transcription factors across the cell cycle. bioRxiv 2019. doi:10.1101/698571.
Professor David Suter
Professor David Suter studied medicine at the University of Geneva in Switzerland, where he obtained a MD/PhD in the field of stem cell biology. After gaining postdoctoral experience in single cell and single molecule analysis of gene expression, he established his laboratory at the Bioengineering Institute of EPFL in 2013.
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