We used both unbiased and candidate approaches to identify regulators of nucleolar size and function. First, we performed EMS mutagenesis in C. elegans and screened for mutants with enlarged nucleoli using the nucleolar marker FIB-1::GFP. As expected, we recovered new mutations in ncl-1, which we previously showed expands nucleoli and reduces longevity. We also identified several epigenetic regulators that altered reporter expression but did not affect nucleolar size or aging. We therefore focused on how ncl-1 regulates lifespan by integrating transcriptomic, proteomic, and phenotypic analyses, primarily comparing long-lived glp-1 mutants with short-lived glp-1 ncl-1 double mutants.
We found that mutation of ncl-1 induces accelerated aging, characterized by premature senescence markers and reduced mobility. Molecularly, glp-1 ncl-1 mutants show dysregulated ribosome biogenesis: rRNA transcription in the nucleolus is increased while mature rRNA levels decline. mRNA and protein levels of ribosomal components become uncoupled, disrupting ribosomal stoichiometry, impairing translation efficiency, and increasing protein aggregation (Martinez-Miguel, Popkes-van Oepen et al., 2026, BioRxiv). An unbiased RNAi screen revealed that knockdown of the mitochondrial ribosomal protein mrps-16 and the RNase P/MRP component popl-1 restores ribosomal balance and rescues longevity in glp-1 ncl-1 mutants, identifying key components that can bypass nucleolar dysfunction. Ongoing work investigates how mitochondrial–nucleolar cross-talk links metabolism to aging.
In a candidate approach, we examined the argonaute protein NRDE-3, a nuclear RNAi factor that interacts with NCL-1 in yeast two-hybrid assays. Similar to ncl-1, double mutants of glp-1; nrde-3 are short-lived, although nrde-3 appears to regulate lifespan independently of nucleolar function. Current RNAi screens aim to identify the mechanisms involved.
To assess nucleolar function in mammals, we generated mice with a heterozygous deletion of nucleolar fibrillarin and are conducting integrative systems biology and phenotypic analyses, including lifespan and metabolic studies. In parallel, we showed that nucleolar size may be a viable biomarker of aging and inflammation in mice and humans. Enlarged nucleoli correlate with reduced stem cell potential in hematopoietic stem cells in mice (Lengefeld et al., 2021, Science Advances) and with reproductive senescence in C. elegans germline stem cells (Nonninger, Mak, Gerisch et al., 2025, Nature Aging). In mouse kidney injury models, increased nucleolar size correlates with tissue damage but can be prevented by dietary restriction (Koehler et al., 2022, Translational Research). In human blood samples, lymphocytes from older donors show increased fibrillarin levels. Moreover, nucleoli enlarge during infection and inflammation in clinical cohorts and in patients with periodic fever syndromes (Steiner et al., 2023, Rheumatology). Ongoing longitudinal studies aim to establish nucleolar size as a biomarker of inflammation and disease activity.
