Nucleolar reorganization on stress depends on physicochemical changes due to reduced nascent rRNA synthesis
Research Summary: Under cellular stress, nucleolar reorganization naturally stems from decreased nascent rRNA synthesis and subsequent biophysical restructuring, rather than active signaling to mediate rDNA repair.
Researcher Spotlight
Sinjini Ghosh is a researcher at TIFR Hyderabad investigating cellular responses to genomic stress. Her work focuses on nucleolar mechanics and the spatial-temporal reorganization of sub-organelles during DNA damage repair.
Lab PI name: Aprotim Mazumder
University: TATA Institute of Fundamental Research, Hyderabad
Lab social media: https://www.tifrh.res.in/~aprotim/people/
What was the core problem you aimed to solve with this research?
The core problem we set out to address is resolving the exact mechanistic driver behind how the cell nucleolus reorganizes during cellular stress. Historically, it has been well documented that cellular stresses—such as DNA damage or RNA Polymerase I (RNAPI) inhibition—cause the nucleolus to drastically alter its structure (such as forming nucleolar caps or coalescing sub-compartments).
However, two major challenges still existed in the field:
- Resolution Constraints: Standard optical diffraction limits made it difficult to resolve the fine, nanoscale architecture of sub-nucleolar domains and map specific RNA transcripts within them.
- Mechanistic Ambiguity: It remained unclear whether nucleolar reorganization was an active, signal-mediated downstream pathway to facilitate rDNA repair or a passive consequence of disrupted transcription.
How did you go about solving this problem?
We combined super-resolution by optical realignment (So-Ra) and single-molecule fluorescence in situ hybridization (smFISH). This high-resolution approach allowed us to directly visualize sub-nucleolar domains alongside nascent and mature rRNA transcripts during stress conditions (such as DNA damage or RNAPI inhibition). We also used expansion microscopy to visualize sub-nucleolar domains at nanoscale resolution.
To complement these spatial mapping techniques with functional dynamics, we used fluorescence recovery after photobleaching (FRAP) to measure the internal liquid-like dynamics and phase-separation behavior of key nucleolar proteins during stress. Additionally, we employed fluorescence anisotropy to assess local micro-viscosity and structural order within distinct sub-nucleolar phases, providing a comprehensive view of how stress induced reduction in nascent rRNA alters both the spatial architecture and biophysical properties of the nucleolus.
“Sinjini has done a commendable job showing nucleolar reorganization under stress arises naturally by tuning nascent rRNA levels, which the cell in turn can exploit to functional ends. This is an important study from both a scientific and a methodological standpoint.” – Aprotim Mazumder
How would you explain your research outcomes (Key findings) to the non-scientific community?
Imagine the nucleolus as a busy factory assembly line, where freshly made RNA molecules act like onsite project managers. Their physical presence keeps all the different assembly departments connected and running smoothly side-by-side.
When the cell experiences stress, production stops and new project managers disappear. Without them, interdepartmental connections are lost. Isolated from each other, similar workers naturally bundle together into tightly packed groups, turning a flexible factory floor into a frozen, solid-state system—not because anyone ordered a reorganization, but simply because the people keeping the floor connected were gone.
What are the potential implications of your findings for the field and society?
These findings reframe nucleolar stress response as a biophysical consequence of rRNA synthesis rather than a pure signaling cascade. This provides a deeper foundation for understanding phase separation in organelles, which could aid in understanding nucleolar dysfunction and aberrant ribosome biogenesis, associated with diseases like cancers and neurodegenerative disorders.
What was the exciting moment during your research?
Initially when we tried using expansion microscopy to visualize nucleolar compartments, one day during imaging, I observed the ring-like structures in one of the channels that I stained for- it was UBF1 (FC compartment protein). It looked really beautiful and felt great that we were able to gain that resolution. On a different incident, when we visualized nascent rRNA, I saw that upon damage its levels are not completely vanishing as we had expected, but are lower than control, with the remaining rRNA forming a distinct localization moving toward the nucleolar periphery forming caps. It was a new observation with direct probing of nascent rRNA upon damage, and I felt it was quite cool!
Paper reference: https://link.springer.com/article/10.1038/s44319-026-00932-z
