# Genome-wide Investigation of cis-splicing between Adjacent Genes

> **NIH NIH R01** · UNIVERSITY OF VIRGINIA · 2021 · $323,000

## Abstract

PROJECT SUMMARY
Genes and their products (RNA and protein) are not expected to intermingle, except in the situation of cancer.
However, this traditional dogma is being challenged when more and more chimeric RNAs being identified in
non-cancer samples. Our work on trans-splicing and cis-splicing between adjacent genes (cis-SAGe) have
provided evidence that the intergenic splicings may be another mechanism to expand functional genome. In
this proposal, we are focusing on the genome-wide study of the transcriptional read-through fusions (we
named them cis-SAGe to differentiate from other mechanisms). Traditionally, they were believed to be rare, or
artifacts. Even when proven true, they were thought to be transcriptional noise or side products. However, in
our preliminary studies, we have identified thousands of such fusion RNAs in non-cancer tissues and cells.
Some have been validated, and proven functional. We hypothesize that cis-SAGe chimeric fusion RNAs are a
widely spread phenomenon in normal physiology, and represent a means to diversify our transcriptome. To
test this hypothesis, and to gain knowledge about these fusions at the genome level, we propose the following
four aims: In Aim1, we will characterize the fusion RNAs and parental genes including junction sequence,
protein-coding potential, expression, and gene ontology. The fusion RNAs will also be analyzed at multiple
levels with multiple techniques including non RT-based Nanostring platform, and high throughput mass
spectrometry. In Aim2, we will investigate the biological significance of the cis-SAGe fusions. A high throughput
screening will be used followed by candidate approaches with both gain- and loss-of-function systems. For the
loss-of-function system, we will use RNAi method, which is more mature in our hands. In addition, we will use
our newly adapted dCAS9-KRAB method to achieve fusion-specific silencing. We will apply the perturbation in
both cell culture and animal systems. One of the fusions will be examined in more detail in a “retrogenic” mice
model. In Aim3, we will investigate the generating mechanisms of cis-SAGe with two approaches, a
bioinformatics approach and a reporter approach. Bioinformatically, we will interrogate multi-omics data from
ENCODE and Roadmap Epigenomics to investigate epigenetic signatures, transcriptional regulatory patterns,
DNA methylation as well as the three-dimensional proximity of the cis-SAGe fusion sites. In the reporter
system, we have built a two-part renilla luciferase separated by introns, exons and termination site of a model
fusion RNA, and we are using the system to screen candidate factors. In Aim4, we will develop an interactive
web-based database to allow end users to search for fusion RNAs in normal tissues and cells. The findings
from the proposed study will have the potential of not only challenging traditional dogmas that chimeric RNAs
are cancer-specific features, but also enhancing our understanding of the human genome a...

## Key facts

- **NIH application ID:** 10217201
- **Project number:** 5R01GM132138-03
- **Recipient organization:** UNIVERSITY OF VIRGINIA
- **Principal Investigator:** HUI LI
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $323,000
- **Award type:** 5
- **Project period:** 2019-09-03 → 2023-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10217201

## Citation

> US National Institutes of Health, RePORTER application 10217201, Genome-wide Investigation of cis-splicing between Adjacent Genes (5R01GM132138-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10217201. Licensed CC0.

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