Reconstitution of minimal nucleoli in vitro – characterization of protein/rRNA interactions and LLPS properties

https://iab-grenoble.fr/fr/l-institut

Synopsis: By combining fluorescence imaging with molecular and cell biology techniques the objective of the internship is to characterize physicochemical parameters that drive liquid-liquid phase separation of nucleolar proteins with rRNA.

Abstract: Join the international young and dynamic team working with state-of-the-art experimental approaches at the interface of cell biology and mechanobiology. 

Our research identifies pathways of signaling enabling cells to rapidly adapt to different environmental stresses. We recently uncovered that nucleolus – a molecular biocondensate - is a mechanosensitive structure that remodels upon osmotic and mechanical stress (doi.org/10.64898/2026.07.02.731374; under review in Nature Cell Biology). The adaptation process is associated with the translocation of nucleolin. However, proteomic analysis of its PTMs did not show any regulation at this level suggesting alternative mechanisms triggering this nucleolin shift. Our current hypothesis is that mechanical stress induces changes in the physico-chemical environment (crowding, pH, ionic force) that impact the condensate stability and partition coefficient of nucleolin. To test this hypothesis, we elaborated protocol of full-length nucleolin purification and characterized LLPS (liquid-liquid phase separation) phase diagrams of two abundant nucleolar proteins: nucleolin and nucleophosmin. For the proposed internship we would like the candidate to characterize the LLPS of this bi-composite system with addition of ribosomal RNA. These experiments will reveal how physico-chemical environment triggered by mechanical stress regulate nucleolin-based signaling. The subject has a potential to develop into a PhD thesis proposition after M2 with extension to test the observed in vitro phenomena in an in vivo model of Xenopus oocytes. 

Methods: Candidate will use state of the art high-resolution microscopy to observe phase separation and using developed analysis pipeline will perform quantitative analysis of condensates size, enrichment, and rheology (FRAP) to determine phase diagrams of the compositive system. 

Selected publications of the team relevant to the subject: 

1) Shetty Y., Elias K.O., Badawi S., Pernet L., Ribba A.-S., Oddou C., Wacheul L., Belmudes L., Moutaux E., Zorbas C., Fraboulet S., Couté Y., Erdel F., Lafontaine D.L.J., Dolega M.E. The nucleolus is a mechanosensitive condensate that adapts ribosome biogenesis to mechanical forces, 2026.07.02.731374, bioRxiv, 2026, (under review at Nature Cell Biology).

2) Shetty Y. and Dolega M.E.*, Mechanobiology of the nucleolus. Biology of the Cell 118 (2), e70052, 2026.

https://iab-grenoble.fr/fr/recherche/equipe/axes/dynamique-du-cytosquelette-et-fonctions-nucleaires#64

Motivated to work in a highly interdisciplinary team. Experience in fluorescence microscopy and basic cell biology techniques is a plus.