Nucleolar Stress and Disease
Our research aims to understand how misfunctioning of a specific part of human cells, an organelle called the nucleolus, contributes to cancer development and how this process can be targeted to offer novel treatment opportunities for cancer patients.
Our research
The nucleolus is essential for protein production and regulation of cell growth. High nucleolar activity in cancerous cells supports rapid cell growth, and inhibition of nucleolar activity can induce cell death in cancer cells. The nucleolus therefore offers an attractive target for development of novel anti-cancer treatments.
A particular subset of genes, the ribosomal RNA genes (rDNA), are found only in the nucleolus. Due to the high cellular demand for protein, particularly in rapidly growing cells, the rDNA is the most active genomic region and each cell is estimated to have around 400 copies of the ribosomal RNA genes. The rDNA is an intrinsically unstable region of the genome and one of the most frequently rearranged parts of the genome in cancers.
The Nucleolar stress and disease group aims to reveal the molecular mechanisms that maintain and protect the rDNA. The study of DNA damage in specific genomic regions has been technically challenging in the past, but the discovery of CRISPR/Cas9 technology provided new opportunities as it allows site-specific induction of DNA breaks, a technique that is used and continuously developed in the lab (Korsholm et al., 2019). By introducing guide RNAs targeting rDNA, we can create nucleolar DNA damage recognized by NBS1-GFP and we can quantify rDNA repair by measuring GFP-intensity in the nucleolus (See Figure 1A and B)
One of our aims is to uncover novel pathways protecting rDNA integrity. For this purpose we use high-throughput imaging screens, proximity labelling techniques and mass spectrometry analysis. We analyse the consequences of defects in the nucleolar DNA damage response, such as changes in cell growth, survival, ribosome biogenesis and genome instability, with a range of biochemical techniques, high-content imaging and superresolution microscopy. We also investigate how inhibition of newly identified rDNA repair proteins can be used for cancer treatment.
A second aim is to develop nucleolar biomarkers to improve treatment of cancer patients. We are developing methods to assess nucleolar status in cancer patients to further understand how increased nucleolar activity promotes cancer development, can be targeted, and influence the outcome of currently used cancer treatments.
Find us at X: Larsen_Lab
Ongoing projects
The nucleolar DNA damage response
The nucleolar DNA damage response is unique and in addition to break detection, signalling and repair it also involves transcriptional silencing and a prominent restructuring of the nucleolus that leads to accumulation of rDNA at the nucleolar periphery in so-called nucleolar caps (See Figure 1B).
We have conducted high-throughput screens (using siRNA libraries and proximity-based labelling techniques) to identify new proteins that control this process. We are currently characterizing candidate proteins to determine their individual roles in the nDDR and to determine their anti-cancer potential.
Mutations in several well-characterized DDR proteins lead to hyper-instability of rDNA. By depletion and knock out of such DDR proteins, we also study mechanisms maintaining rDNA stability and the faulty mechanisms that become activated in their absence and lead to rDNA destabilization.
The nucleolus and cancer
Nucleolar activity is upregulated in almost all cancers and altered nucleolar morphology is a characteristic of cancer. Despite this our understanding of nucleolar dysregulation in cancer and its clinical use is very limited. We aim to establish methods that will enable evaluation of nucleolar status in cancer patients and thereby allow new investigation of its predictive and/or prognostic value.
Oxe KC, Larsen DH. Treacle is Upregulated in Cancer and Correlates With Poor Prognosis. Front Cell Dev Biol. 2022 Jun 20;10:918544. doi: 10.3389/fcell.2022.918544. PMID: 35794866; PMCID: PMC9251355.
Gál Z, Nieto B, Boukoura S, Rasmussen AV, Larsen DH. Treacle Sticks the Nucleolar Responses to DNA Damage Together. Front Cell Dev Biol. 2022 May 12;10:892006. doi: 10.3389/fcell.2022.892006. PMID: 35646927; PMCID: PMC9133508.
Korsholm L, Gal Z, Nieto B, Quevedo Oliver, Boukoura S, Lund C, Larsen DH. Recent advances in the nucleolar responses to DNA double-strand breaks. Nucleic Acids Res. 2020 Sep 25;48(17):9449-9461. doi: 10.1093/nar/gkaa713.
Korsholm LM, Gál Z, Lin L, Quevedo O, Ahmad DA, Dulina E, Luo Y, Bartek J, Larsen DH. Double-strand breaks in ribosomal RNA genes activate a distinct signaling and chromatin response to facilitate nucleolar restructuring and repair. Nucleic Acids Res. 2019 Sep 5;47(15):8019-8035. doi: 10.1093/nar/gkz518. PubMed PMID: 31184714; PubMed Central PMCID: PMC6735822.
Larsen DH, Hari F, Clapperton JA, Gwerder M, Gutsche K, Altmeyer M, Jungmichel S, Toledo LI, Fink D, Rask MB, Grøfte M, Lukas C, Nielsen ML, Smerdon SJ, Lukas J, Stucki M. The NBS1-Treacle complex controls ribosomal RNA transcription in response to DNA damage. Nat Cell Biol. 2014 Aug;16(8):792-803. doi:10.1038/ncb3007. Epub 2014 Jul 27. PubMed PMID: 25064736; PubMed Central PMCID: PMC4962910.
Group leader: Dorthe Payne-Larsen
Dorthe Helena Larsen leads the Nucleolar Stress and Disease Group. She has a strong background in molecular biology with a deep interest in the cellular response to DNA damage.
Dr. Larsen received her PhD degree from the University of Copenhagen in 2009 for her research conducted at the Danish Cancer Society. During her PhD-studies she worked in the laboratory of Professor Jiri Lukas and Professor Jiri Bartek on identification of novel DNA repair proteins. Dr. Larsen continued her postdoctoral studies at the University of Zurich, Switzerland, in the lab of Professor Manuel Stucki and as a visiting researcher at the Novo Nordisk Center for Protein Research, Denmark, where she identified a protein complex that regulates nucleolar function after DNA damage.
In 2016 Dr. Larsen started the Nucleolar Stress and Disease Group at the Danish Cancer Society that focuses on DNA repair of the ribosomal RNA genes. In 2018 she received the prestigious starting grant Sapere Aude from the Independent Research Fund Denmark.
ORCID: 0000-0002-8280-1252
Key funding
The Danish Cancer Society
Knæk Cancer
Sapere Aude Starting Grant, Independent Research Fund Denmark
The Novo Nordisk Foundation.