# Nuclear speckle liquid-liquid phase separation dynamics in senescence and aging

> **NIH NIH F31** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2024 · $48,974

## Abstract

Abstract
Organismal health requires a consistent and balanced internal environment known as homeostasis. Different
physiological processes maintain proper levels of biomolecules at a cellular level, and several of these
mechanisms lose efficacy with age. Proteostasis, sustained levels of correctly folded proteins in the endoplasmic
reticulum (ER), is maintained by the Unfolded Protein Response (UPR). Excessive misfolded proteins in the ER
activate the three branches of the UPR, facilitating adaptive processes to restore a balanced proteome in the
cell. Aging is associated with the loss of proteostasis and the accumulation of senescent cells – cells that no
longer replicate and secrete pro-inflammatory signals – that exhibit a dysfunctional UPR. The molecular
mechanisms underlying the altered UPR in senescent cells are unclear. We hypothesize that the liquid-liquid
phase separation (LLPS) dynamics of a nuclear biomolecular condensate, the nuclear speckle (NS), link cellular
senescence to the UPR. The 12-hour, XBP1s-dependent clock that functions independently of the 24-hour clock
or the cell cycle establishes 12-hour ultradian rhythms of NS LLPS dynamics. These rhythms regulate NS
morphology and fluidity through SON, the NS core protein. High SON levels create a diffuse, fluid NS and boost
the expression of UPR-associated genes. In contrast, low SON levels result in a spherical, stagnant NS and a
blunted expression of UPR genes. We have recently found that SON levels decrease, and that the NS becomes
more spherical during cellular senescence. These data suggest that changes to NS LLPS dynamics are
hallmarks of cellular senescence and aging. Here, we propose two aims to examine how NS LLPS dynamics
change in vitro during cellular senescence and in vivo throughout chronological aging. In the first aim, we will
use a mouse embryonic fibroblast line with a GFP-tagged NS that can be induced to enter senescence. This
model will examine how established 12-hour rhythms of NS LLPS dynamics change during senescence and how
restoring SON levels affects NS LLPS dynamics in senescent cells. We will also pharmacologically boost the
diffuseness of the NS to determine if its fluidity can be increased during senescence. The second aim will use a
Caenorhabditis elegans (C. elegans) model with a GFP-tagged NS. We will examine how NS LLPS dynamics
change throughout aging and if genetic and pharmacological methods that make the NS more diffuse in
mammalian cells can similarly affect NS LLPS dynamics in C. elegans and boost proteostasis in aged organisms.
These aims will establish changes to NS LLPS dynamics as hallmarks of senescence and aging. Furthermore,
we intend to show that NS LLPS dynamics is a druggable target and that therapies could return the NS
morphology and fluidity to a pre-senescent state.

## Key facts

- **NIH application ID:** 10846553
- **Project number:** 5F31AG080998-02
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** William Aaron Dion
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $48,974
- **Award type:** 5
- **Project period:** 2023-04-01 → 2026-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10846553

## Citation

> US National Institutes of Health, RePORTER application 10846553, Nuclear speckle liquid-liquid phase separation dynamics in senescence and aging (5F31AG080998-02). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10846553. Licensed CC0.

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