Team FABRE & LESAGE

Biology of Genome Instability (GeBi)

Double-strand breaks represent the most severe form of DNA damage a cell can sustain. Left unrepaired, they can cause chromosomal abnormalities such as aneuploidy, genetic mutations, or even cell death. Homologous recombination is one of the universal mechanisms for correcting these lesions. It uses an intact DNA molecule as a template to restore genome integrity. The first phase of this process relies on homology search and DNA strand invasion, a step catalyzed by the Rad51 protein complex. However, the complexity of this initial step is compounded by the spatial organization of chromosomes within the nucleus, as well as by the presence of repeated sequences, such as tandem satellite DNA, telomeric DNA, and transposable elements. These features complicate our understanding of this critical step in double-strand break repair.

Our goal is to determine how chromosome organization influences homology search during the repair of a double strand break within unique and repeated sequences such as Ty2 retrotransposons and telomeric sequences. This research axis relies on live-cell time-lapse microscopy combined with molecular and cell biology approaches.

The contribution of retrotransposon-type transposable elements to cancer development is generally associated with their mutagenic potential and their ability to deregulate gene expression. However, recent studies reveal that intermediate steps of retrotransposition can also have significant effects on health, underscoring the importance of understanding the regulation of all steps of retrotransposon replication. Retrotransposon RNA is essential to their replication, serving as a template for both translation and reverse transcription. Its properties, as well as its interactions with cellular proteins, remain poorly understood.

Our goal is to characterize retrotransposon ribonucleoparticles (RNPs) using the S. cerevisiae Ty1 retrotransposon as a model. These proteins could regulate the fate of retrotransposon RNAs, modulate retrotransposition, and contribute to genome protection. This research axis relies on the development of an innovative proteomic approach combined with molecular and cell biology methods.

Retrotransposons, such as Ty1 in yeast, are major mobile elements of eukaryotic genomes. Their integration into DNA depends on a complex chromatin environment, where accessibility is regulated by DNA-binding proteins, chromatin remodelers, and histone modifications. However, the precise mechanisms guiding their integration remain poorly understood.

We have previously demonstrated that Ty1 integrates preferentially upstream of genes transcribed by RNA polymerase III (Pol III), through an interaction between its integrase and Pol III (Bridier-Nahmias, Science 2015; Asif-Laidin, EMBO J 2020). In the presence of the integrase, Pol III adopts a distinct conformation, extending its residence time on chromatin and promoting Ty1 integration upstream of Pol III-transcribed genes (Nguyen, Nature Communications, 2023). When the interaction with Pol III is abolished, Ty1 targets subtelomeres, suggesting a role for local cofactors or chromatin marks in this process.

Our goal is to understand how chromatin dynamics and genome organization guide retrotransposon integration — a model for retroviruses. To identify the interactions between Ty1 integrase, its cofactors, and chromatin that are essential for successful integration, we combine multiple approaches including molecular and structural biology, genetics, and genomics.