The field has well established that mutations in SF3B1, U2AF1, SRSF2, and ZRSR2 are recurrent drivers in cancers, especially myeloid malignancies, causing aberrant isoform switching. Concurrently, a new biophysical dimension has emerged: liquid-liquid phase separation (LLPS) dysregulation by mutant splicing factors can sequester splice sites, promoting pro-tumorigenic isoforms.

Yet, the overlapping role of splicing in aging—where altered expression of mRNA splicing factors and deregulation of alternative splicing are hallmarks of aging that contribute to cellular senescence—remains a mechanistic paradox. How does a single biochemical network drive both hyper-proliferative oncogenesis and anti-proliferative senescence? Furthermore, whether shared splicing defects causally link accelerated aging to increased cancer risk remains unknown.

The Hypothesis

I propose that the age-dependent decline in nuclear ATP acts as a biophysical switch that dictates the phase state of spliceosomal condensates. ATP is a well-established biological hydrotrope that maintains the solubility of intrinsically disordered proteins. I hypothesize that as nuclear ATP concentrations fall during aging, splicing factor condensates undergo a pathological liquid-to-solid phase transition (gelation), which alters their functional specificity.

Mechanistic Divergence: Senescence vs. Oncogenesis

This biophysical shift creates a bifurcation in cellular fate depending on the mutational landscape:

1. In wild-type aging cells (Senescence): Splicing factors such as SRSF1—which has been identified as a rejuvenation factor—require high ATP levels to remain fluid. As ATP drops, SRSF1 becomes trapped in rigid, gel-like nucleoplasmic condensates. This functionally depletes the soluble pool of SRSF1, driving global intron retention, stalling the cell cycle, and inducing senescence. This explains why the functional effects of splicing factors can be highly context-dependent; their behavior is governed by the local metabolic/hydrotropic environment.

2. In aging cells with acquired mutations (Oncogenesis): Mutations like SRSF2^P95H fundamentally alter the protein's LLPS valency and RNA-binding affinity. While mutant SRSF2 enriches U1/U2AF condensates, causing exon skipping in genes like BRCA1 even in healthy cells, the age-related drop in ATP causes these specific mutant condensates to hyper-assemble into highly stable, aberrant processing hubs. Instead of inducing global splicing failure (senescence), these solid-like mutant condensates selectively sequester and mis-splice targeted tumor-suppressor pre-mRNAs, actively driving oncogenesis.

Falsifiability & Experimental Design

This hypothesis shifts the focus from purely transcriptomic changes to the biophysical material properties of the nucleus. It can be directly tested via:

• In Vitro LLPS Assays: Reconstitute wild-type SRSF1 and mutant SRSF2 condensates and titrate physiological vs. pathological ATP concentrations. Prediction: WT SRSF1 will show a sharp liquid-to-solid transition at low ATP, whereas mutant SRSF2 will form precipitous, irreversible gels.
• In Vivo FRAP (Fluorescence Recovery After Photobleaching): Measure the mobility of SRSF1 and SRSF2 in young vs. senescent fibroblasts. Prediction: Senescent cells will show significantly slower recovery rates (gelation), which should be reversible upon exogenous ATP supplementation or hydrotrope administration.
• Therapeutic Rescue: The observation that small molecules can modulate splicing factor expression and reverse multiple features of cellular senescence might actually reflect these molecules acting as chemical hydrotropes that dissolve solid-phase condensates.

By framing splicing dysregulation as a thermodynamic problem driven by nuclear metabolomics, we provide a unified mechanistic basis for why aging promotes both senescence and a permissive environment for splicing-driven oncogenesis. This insight could fundamentally alter how we design next-generation spliceosome modulators and antisense oligonucleotides (ASOs).