Switch it off: new insights into repressors of gene transcription
Loni Klaus joined the Vienna BioCenter PhD Program in 2018 to study the regulation of gene transcription in the lab of Alexander Stark. She recently submitted her doctoral thesis and published her findings in The EMBO Journal. In this interview, she tells us more about her research.
Why is the regulation of gene transcription important for multicellular organisms?
Almost all cells in an organism contain the same genetic information, but each cell type only transcribes a subset of genes. Regulating transcription is crucial to determining a cell’s shape and function in the body. A muscle cell, for example, produces proteins that give it the ability to contract, while a neuron makes proteins that enable it to transmit signals. Which genes are transcribed, when, and at what level plays a big role in shaping a cell’s identity.
What approach did you take to study gene regulation?
Simply put, the regulation of gene transcription is a two-sided process: genes can be activated or repressed. Most labs in the field have investigated how genes get activated – how and when do genes become active to influence what goes on in a cell? The other side has received much less attention until recently, and that is what I wanted to address in my PhD. How can a gene be turned off? This question is extremely important for cell identity – a neuron should not produce proteins that make it contract like a muscle cell, to stick to my example.
What are open questions in this field?
Repressive transcription factors bind to DNA motifs in the regulatory sequence of their target gene, and they typically recruit other molecules, called co-repressors, that turn off the expression of this gene. There are relatively few studies on such transcriptional repressors and most of them consider a handful of proteins of interest. I was more interested in studying them from a systems biology point of view, meaning that I wanted to learn more about transcriptional repressors as a whole and differences among them. What are all these repressive transcription factors? What do the co-repressors look like? Which of them interact, and how do they mediate repression? So many open questions!
How did you catalogue repressive transcription factors?
These proteins have two domains: the DNA-binding domain, and the repressive domain that recruits co-repressors. In my project, I developed a method called RD-seq (for repressive-domain-sequencing) and used it on fruit fly S2 cells with an integrated transcriptional reporter: I screened all transcription-related proteins and identified 195 repressive domains. One of the goals of my projects was to figure out if these domains can be organised in functional subgroups, based on their sequence of amino acids.
What did you find then?
We found specific sequences or ‘motifs’ of only a few amino acids that appear in many repressive domains, suggesting that they are important for function. When I experimentally mutated five of these motifs, the repressive domains couldn’t function anymore. Another key finding is that repressive domains with these motifs interact with specific co-repressors – the presence of such a motif can help us predict which co-repressor will be recruited.
Do these findings apply to other organisms?
Yes, we showed that most of the motifs that are important for the function of repressive domains are conserved in animals, from fly all the way to humans. This gives us the chance to predict potential repressive domains in human transcription factors based on the presence of these motifs.
Where do you see this project going next?
I think the list of repressive domains, their repressive motifs and interacting co-repressors we generated is a great resource to better understand repression of gene transcription and the impact of mutations within repressive transcription factors. As a next step, it would be interesting to investigate how exactly genes are shut down. Little is known about repressive mechanisms, and I’d be curious to find out if they differ between co-repressors. With this information, we could build a “catalogue of repression” containing repressive transcription factors, their repressive domains and motifs, the associated co-repressors, and repressive mechanism.
Are there applied contexts in which such a catalogue could come in handy?
There are several genetic disorders linked with mutations in transcriptional repressors. One example is Rett syndrome, which is a neurodevelopmental disorder that can be caused by a mutation in a repressive domain. The mutation prevents the interaction with the co-repressor and messes with normal gene transcription. Cataloguing repressive transcription factors and their co-repressors could help us explain disease mechanisms and potentially find a way to counter them.
Loni Klaus, Bernardo P. de Almeida, Anna Vlasova, Filip Nemcko, Alexander Schleiffer, Katharina Bergauer, Lorena Hofbauer, Martina Rath, Alexander Stark: “Systematic identification and characterization of repressive domains in Drosophila transcription factors”. The EMBO Journal (2022), DOI: 10.15252/embj.2022112100.