Projektbeschreibung
Regulierung der Torsionsspannung der DNA auf der Kernlamina
Bei der DNA-Transkription oder -Replikation kommt es zu einer Torsionsspannung in der Doppelhelix. Das Enzym, das diese Spannung beseitigt, ist DNA-Topoisomerase I (Top1), ein Ziel der Krebs-Chemotherapie. Ein Verlust der Aktivität von Top1 führt zu einer Akkumulation der Torsionsspannung in transkriptionell aktiven Genen, wodurch sich nichtkanonische DNA-RNA-Hybridstrukturen bilden, die R-Schleifen genannt werden. Kürzlich konnte das Projektteam zeigen, dass der Verlust von Top1 zu einer Stabilisierung der R-Schleifen führt, vor allem in Genen, die auf der Kernlamina verankert sind. Diese Beobachtung deutet darauf hin, dass sich die Dissipation der Torsionsspannung durch das Anhaften der DNA an die Kernlamina verhindern lassen könnte. Das EU-finanzierte Projekt TorsionAtLamina untersucht anhand von im Gastlabor entwickelten Geomtechniken den Zusammenhang zwischen Torsionsspannung, Top1, R-Schleifen und der Anhaftung an die Kernlamina.
Ziel
When DNA is transcribed or replicated, torsional stress accumulates on the double helix. This tension must be dissipated by spreading it along the DNA fiber, or it must be removed altogether. One of the main factors responsible for the removal of such stress is DNA Topoisomerase I (Top1), an important target of cancer chemotherapy. When Top1 activity is lost, torsional stress accumulates on transcriptionally active genes and can lead to the formation of non-canonical DNA/RNA hybrid structures called R loops. These structures are emerging as important regulators of genome function and stability. By genome-wide mapping of R loops in human cells, I recently found that depletion of Top1 leads to a marked R loop stabilization, specifically on genes that are anchored to the nuclear lamina. This strongly suggests that attachment of DNA to the nuclear lamina may prevent dissipation of torsional stress, but how this works is still largely unclear. I propose to investigate the causal relationships between torsonal stress, Top1, R-loops and nuclear lamina attachment, taking advantage of a suite of unique genomics techniques developed in the host lab. Specifically, I will: 1) Develop two novel reporter assays to probe the effects of chromatin context (in particular lamina associated chromatin) and Top1 on torsional stress and R loop formation, at thousands of locations in the human genome. 2) Investigate if and how Top1 regulates DNA/nuclear lamina contacts by by means of a novel version of the powerful genome-wide DamID mapping method with much-improved time resolution. My expertise in Top1, R-loops and DNA topology combined with the unique genomics methodologies in the host lab, as well as their expertise in lamina-associated DNA, will lead to a unique synergy that should result in new insights into the relationship between nuclear organization, torsional stress and R loop formation. Moreover, it will yield new methods that will boost scientific progress in this field.
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MSCA-IF - Marie Skłodowska-Curie Individual Fellowships (IF)Koordinator
1066 CX Amsterdam
Niederlande