# Regulation and consequences of cryptic splicing

> **NIH NIH R35** · BAYLOR COLLEGE OF MEDICINE · 2024 · $469,923

## Abstract

ABSTRACT
Our research aims to comprehensively understand the molecular mechanisms and functions of RNA splicing in
regulating cellular phenotypes. RNA splicing is a critical process that removes introns from pre-mRNA and is
essential to gene regulation and transcriptome diversity. In humans, introns constitute approximately one-fourth
of the whole genome, representing a vast expansion compared to lower eukaryotes and other mammals. Long
introns contain a high number of pseudo splice sites that closely resemble those of coding exons. Misuse of
pseudo splice sites gives rise to cryptic splicing, which leads to the erroneous insertion of intronic elements into
mRNA. Consequently, cryptic splicing results in disruption of transcriptome integrity and is associated with
numerous pathologies including neuronal degeneration and cancer. Our proposed work aims to build on our
recent discoveries in investigating the regulatory mechanisms of cryptic splicing and its consequences. We
recently conducted a survey of RNA binding protein (RBP) interactions with introns and identified a group of
RBPs, including heterogenous nuclear ribonucleoprotein M (hnRNPM), whose binding is highly enriched in
introns. We found that depletion of hnRNPM by shRNA causes over a thousand cryptic splicing events,
suggesting that hnRNPM is a crucial repressor of cryptic splicing. Our bioinformatics analysis shows that
hnRNPM binding is enriched in regions that contain repetitive sequences. It further predicts that hnRNPM-
repressed cryptic splicing products can be targets of nonsense-mediated decay or templates to generate
truncated proteins. The cryptic splicing products also have the potential to form double-stranded RNAs (dsRNAs)
that trigger an interferon response, leading to significant increases in the expression of interferon-stimulated
genes. Building on these findings, we will first comprehensively investigate the underlying mechanisms by which
RBPs repress cryptic splicing to maintain transcriptome integrity. We will investigate the role of RBPs that
preferentially bind to introns, including hnRNPM interaction partners, in repressing cryptic splicing. We will also
delineate the nature of interactions between these RBPs and hnRNPM. Furthermore, we will determine the
molecular details of how loss of hnRNPM affects spliceosome factor binding at pseudo splice sites, resulting in
cryptic splicing. Second, we will investigate the fate of cryptic splicing transcripts by experimentally characterizing
them at both the mRNA and protein levels and evaluating their impact on cellular activities. Given our finding
that hnRNPM depletion leads to a dsRNA-dependent upregulation of interferon-stimulated genes, we plan to
further investigate the pathways mediating the crosstalk between dsRNA of cryptic transcripts and upregulation
of interferon-stimulated genes and their impact on cell fitness. Successful completion of these studies will shed
light on the regulatory mechanisms for cryptic sp...

## Key facts

- **NIH application ID:** 10842610
- **Project number:** 2R35GM131876-06
- **Recipient organization:** BAYLOR COLLEGE OF MEDICINE
- **Principal Investigator:** Chonghui Cheng
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $469,923
- **Award type:** 2
- **Project period:** 2019-04-01 → 2029-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10842610

## Citation

> US National Institutes of Health, RePORTER application 10842610, Regulation and consequences of cryptic splicing (2R35GM131876-06). Retrieved via AI Analytics 2026-05-27 from https://api.ai-analytics.org/grant/nih/10842610. Licensed CC0.

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