# Functional consequences of mutations in spliceosomal small nuclear RNAs

> **NIH NIH R01** · CLEVELAND CLINIC LERNER COM-CWRU · 2022 · $472,491

## Abstract

Project Summary
The removal of introns from pre-mRNA transcripts is an essential step in the expression of almost all human
genes. The goal of this project is to determine the molecular, cellular and tissue-specific consequences of
mutations in the U4atac snRNA gene (RNU4ATAC) that are the cause of the rare and severe human
developmental disorders microcephalic osteodysplastic primordial dwarfism type I (MOPD I), Roifman
Syndrome (RS) and Lowry-Wood Syndrome (LWS). U4atac snRNA is required for the splicing of minor class
U12- dependent introns. Genes containing these introns are poorly spliced in MOPD I and RS patient cells. We
hypothesize that the various pathologies seen in MOPD I, RS and LWS are caused by reduced splicing or mis-
splicing of a small number of genes containing U12-dependent introns. In addition, we have found that the
RNU4ATAC gene, as well as other snRNA genes, harbor large numbers of low frequency single nucleotide
polymorphisms in all human populations, supplying a mutational origin for these recessive disorders. We have
developed cell lines, including patient-derived and gene-edited human iPS cells, and new assay systems to
investigate the specific defect(s) in splicing caused by the mutations. 1) We will use these cell lines to analyze
the effects of all possible U4atac snRNA mutations on cell growth using a high-throughput method. Based on
these results and the mutations observed in patients with the different syndromes, we will generate cell lines
homozygous for various mutations and employ biochemical techniques to characterize their functional defects.
2) We will determine the differentiation potential of human iPS cells with a variety of disease-linked mutations.
For example, since severe MOPD I is characterized by microcephaly and lissencephaly, we are using MOPD I
patient-derived human iPS cells to generate and study cerebral brain organoids. We have also generated
mouse mutant U4atac models that recapitulate many MOPD I pathologies. They also develop severe diabetes
by two months of age. We will study the cellular phenotypes of the mouse pancreas as the disease develops to
characterize the developmental disorder. 3) We will use RNA-seq and other molecular analyses of human cells
and mouse tissues to relate specific pathologies to defects in splicing of target genes. These targets will be
validated through restoration of gene expression by properly spliced cDNA. Successful completion of these
studies will advance our understanding of spliceosomal splicing, define the molecular causes of human
diseases and determine the role of U12-dependent splicing in gene expression and development.

## Key facts

- **NIH application ID:** 10468151
- **Project number:** 5R01GM133989-04
- **Recipient organization:** CLEVELAND CLINIC LERNER COM-CWRU
- **Principal Investigator:** MAURA L GILLISON
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $472,491
- **Award type:** 5
- **Project period:** 2019-08-01 → 2024-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10468151

## Citation

> US National Institutes of Health, RePORTER application 10468151, Functional consequences of mutations in spliceosomal small nuclear RNAs (5R01GM133989-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10468151. Licensed CC0.

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