Research
Why we want to understand R-loop biology
Precisely controlled gene expression is a fundamental process for the survival of any organism. It is achieved through tight interconnection between transcription, RNA processing, and translation, ensuring both efficiency and fidelity of gene expression. Now it has become clear that RNA molecules are key regulatory elements in this process, raising the importance of the RNA processing step in the gene expression pathway. Importantly, mutations in components regulating RNA processing can lead to global mis-regulation of gene expression and cause human diseases. Research in our lab focuses on understanding the mechanisms governing gene regulation in humans in health and disease conditions.
Function of R-loops in gene regulation
R-loops are formed in all living organisms where they play important roles in regulating gene expression; DNA and histone modifications; generation of antibody diversity; DNA replication; and genome stability. We demonstrated that R-loops are involved in transcriptional termination (Skourti-Stathaki, Mol Cell 2011, Cristini, Cell Rep 2018) and maintenance of genome stability (Cristini, Cell Rep 2018). We are currently investigating the role of R-loops in different aspects of gene expression, employing gene-specific and whole-genome approaches.
R-Loop Proteome
To understand the molecular mechanisms underlying R-loop biology, we recently employed a Mass Spectrometry-based high-throughput affinity purification to define the R-loop-binding proteome in human cells (Cristini, Cell Rep 2018). Among these R-loop interacting proteins, we detected many uncharacterised interactors, including helicases, chromatin and DNA repair factors. Thus R-loop proteome has provided us with a foundation for studies on R-loop regulation. In particular, it helped us to uncover the role of RNA/DNA hybrid helicase DHX9 in maintaining genome stability (Cristini, Cell Rep, 2018), m6A RNA modification in R-loop turn-over (Abakadir, Nat Genetics, 2020) and structure-specific XPG/XPF endonucleases in R-loop processing (Cristini, Cell Rep, 2019). Currently we are exploring the functions of multiple novel R-loop regulators.
Repeat Expansion Diseases
Around 40 human diseases are associated with expanded repeat sequences. Among them are Friedreich ataxia and Fragile X syndrome. Both disorders are associated with the presence of expanded repeats in the non-coding part of the FXN or FMR1 genes, resulting in their transcriptional silencing. However, the molecular mechanism of this process is not well understood. We discovered that expanded repeats of FXN and FMR1 genes form R-loops in patient cells, and these structures drive formation of repressive heterochromatin marks resulting in the host gene silencing (Groh, PLoS Genetics 2014a). Currently we employ multiple molecular and cell biology approaches to uncover the mechanism of R-loop-mediated transcriptional silencing of FXN and FMR1 genes. The findings from this project will be essential for the development of new therapeutic strategies for expansion repeat disorders.
Other R-loop Pathologies
Mis-regulation of R-loops is associated with multiple human diseases, including cancer and neurodegeneration (Kontsantis, Nature Comm 2016, Groh, PLoS Genetics 2014b). However, the exact molecular mechanisms linking R-loops and the onset of these diseases is not fully understood. We investigate the molecular mechanisms underlying the function of pathological R-loops in these diseases, with the aim of utilising this knowledge to design therapeutic interventions to treat these disorders.
