Non-coding RNAs regulatory networks controlling lateral root organogenesis across Brassicaceae – RESOLAT

Long non-coding RNAs (lncRNAs) are now recognized as regulators of gene expression, from chromatin conformation to protein translation, playing a key role in a variety of biological processes. However, their mode of action remains poorly understood, due to the lack of known characteristics that can determine their mechanism of action. Their integration within biological networks remains largely unknown, in particular lateral root development.

The multidisciplinary RESOLAT project aims to understand the impact of lncRNAs on the gene network regulating lateral root development in Brassicaceae. Using time-series RNA-seq data, we propose to reconstruct the gene regulatory network controlling lateral root development in eight Brassicaceae, integrating coding and non-coding genes (small and long). This approach will enable us to identify new interactions and regulatory pathways, with particular emphasis on lncRNAs.

To overcome the challenges associated with the functional characterization of lncRNAs, we propose an approach based on the conservation of their function. Based on lateral root development, a highly conserved process in Angiosperms, we will use conservation of position in the regulatory network as a proxy for conservation of function, i.e. regulation of orthologous genes in different species. By integrating data on genomic conservation (genomic synteny, sequence, secondary structure) and expression conservation during lateral root development, we will seek to better understand the conservation features leading to the functionality of long lncRNAs, including their ability to regulate orthologous genes within Brassicaceae.

Using deep learning approaches based on large language models (LLMs) that will combine sequence and network data, we plan to develop a predictive tool to determine whether a lncRNA acts as a regulator of gene expression level.

Finally, we will use functional approaches in several species to validate the function of conserved lncRNAs in the control of lateral root architecture. First, we will use a semi-high-throughput method derived from Perturb-seq to confirm the regulatory action of conserved lncRNAs in A. thaliana. Based on these results, we will further functionally characterize lncRNAs conserved in several Brassicaceae species to validate their role in controlling lateral root architecture. By modifying the expression dynamics of conserved regulators, we should be able to quantitatively modify root architecture and confirm the regulatory links established by gene network inference.

By comparing different species with a common lateral root organogenesis program, this project will reveal evolutionary conserved and specific mechanisms controlling lateral root development in Brassicaceae, including both wild and cultivated species. It will also enable us to identify functionally conserved lncRNAs and analyze the features underlying their conservation.