We are currently observing a paradigm shift in biogerontology: aging is increasingly defined by a coordinated, pan-tissue loss of RNA splicing fidelity. Recent consensus highlights that RNA Pol II elongation rates impair co-transcriptional splicing, causing a global increase in splicing errors like intron retention across species. While it is clear that mis-splicing drives diseases like Alzheimer's and Parkinson's, a critical mechanistic gap remains: Why does RNA Polymerase II systematically accelerate with age, and how does DNA damage trigger this splicing deterioration?

I propose that transcriptomic drift is not initiated by a primary failure of the spliceosome itself, but is rather a kinetic consequence of global chromatin relaxation driven by the DNA damage response.

The Hypothesis: Kinetic Decoupling via Epigenetic Erosion

I hypothesize that the accumulation of age-associated DNA damage initiates a cascade that fundamentally alters the physical substrate of transcription—chromatin—thereby accelerating RNA Pol II and decoupling it from the splicing machinery.

1. DNA Damage and NAD+ Depletion: As DNA damage accumulates with age, the DNA repair enzyme PARP1 is chronically activated. PARP1 consumes massive amounts of NAD+, leading to systemic intracellular NAD+ depletion. This addresses the question of how DNA damage responses may disrupt splicing.
2. SIRT Inhibition and Chromatin Relaxation: NAD+ depletion severely impairs the activity of sirtuins (specifically SIRT1 and SIRT6), which normally deacetylate histones (e.g., H3K9ac, H3K56ac) to maintain compact chromatin. Loss of sirtuin activity results in global histone hyperacetylation and an "open," relaxed chromatin landscape.
3. Pol II Acceleration: Chromatin compaction normally acts as a physical "speed bump" for RNA Pol II. In the relaxed epigenomic environment of an aged cell, these speed bumps are removed, causing RNA Pol II to travel at an accelerated elongation rate.
4. Transcriptomic Drift: Because splicing is highly co-transcriptional, accelerated Pol II outpaces the recruitment of core splicing factors (like RNP-6 and RBM-39). The transcription machinery passes the 3' splice site before the 5' splice site is fully recognized, systematically generating intron retention and cryptic exon inclusion. This explains how transcriptomic drift involves coordinated dysregulation across tissues[https://www.aging-us.com/news-room/rna-splicing-and-processing-emerge-as-central-features-of-human-aging-across-tissues].

Falsifiability and Experimental Design

This hypothesis provides a purely mechanistic, falsifiable framework to determine if epigenetic state dictates splicing fidelity via elongation kinetics. We can test this through the following experiments:

• Test 1 (Upstream rescue): Restoring the NAD+ pool (via NMN/NR supplementation) or inhibiting PARP1 in aged in vitro human fibroblasts should re-establish baseline histone deacetylation, slow RNA Pol II elongation to youthful rates, and subsequently rescue splicing fidelity without directly targeting the spliceosome.
• Test 2 (Premature induction): Inducing SIRT6 knockout or treating young cells with pan-HDAC inhibitors should prematurely accelerate RNA Pol II and accurately phenocopy the age-associated transcriptomic drift (specifically intron retention patterns).
• Test 3 (Locus-specific uncoupling): Utilizing a CRISPR-dCas9 system fused to an HDAC domain to artificially condense chromatin at specific loci (e.g., the human ortholog of egl-8) in aged cells should locally slow Pol II and restore proper splicing. If successful, this proves that the physical chromatin state, rather than a global defect in splicing factors, dictates local RNA processing.

While genetic modulation of splicing factors rescues senescence[https://doi.org/10.1101/2023.11.13.566787] and antisense oligonucleotides and CRISPR can restore isoform balance, these interventions may be acting downstream. If this hypothesis holds, the "master regulator" of transcriptomic drift is actually the NAD+/SIRT-mediated chromatin landscape. By targeting the epigenome to restore optimal RNA Pol II elongation kinetics, we may be able to universally correct age-associated transcriptomic instability.