# "SHROOM3 is a Novel Cause of Congenital Heart Disease"

> **NIH NIH K08** · INDIANA UNIVERSITY INDIANAPOLIS · 2022 · $155,248

## Abstract

Congenital heart disease (CHD) is the most common cause of death due to birth defects. Despite CHD
frequency, the etiology remains mostly unknown. Understanding CHD pathogenesis will help establish
prognosis, identify comorbidity risks, and develop targeted therapies. We have identified a novel CHD candidate,
SHROOM3. SHROOM3 induces actomyosin constriction and is implicated in cleft palate, neural tube and kidney
defects in humans, but is unexplored in the heart. A recent study demonstrates that SHROOM3 interacts with
Dishevelled2 (DVL2), a component of the noncanonical Wnt/planar cell polarity (PCP) signaling pathway,
suggesting that SHROOM3 may serve as an important link between actomyosin constriction and PCP signaling.
PCP is required for cardiac development, driving neural crest and second heart field migration to the outflow tract.
I utilized a Shroom3 gene-trap mouse (Shroom3gt/gt) to demonstrate SHROOM3-loss-of-function leads to cardiac
defects, including outflow tract defects, phenocopying PCP disruption. Additionally, through whole exome
sequencing in patients with PCP related CHD phenotypes, I identified rare and potentially damaging variants
within SHROOM3's PCP-binding domain. I hypothesize SHROOM3 is a novel component of PCP signaling and
disruption causes CHD. To test this hypothesis, Aim1a assays genetic interaction between SHROOM3 and PCP
during cardiac development, by generating compound heterozygous Shroom3gt/+;Dvl2-/+ embryos and assaying
for a cardiac defects. Aim1b assays Shroom3gt/gt embryos using western blot (WB) for phosphorylated, activated,
PCP components and immunohistochemistry (IHC) analyses for neural crest cell and second heart field migration
to the heart. Aim1c tests the hypothesis that SHROOM3 loss of function disrupts the PCP transcriptional profile
by assaying Shroom3gt/gt hearts using RNA-Seq analysis. Aim 2 tests the hypothesis that specific SHROOM3
variants cause disrupted PCP signaling in vitro. Utilizing CRISPR/Cas9 genome editing, I will generate
SHROOM3 variant induced pluripotent stem cell lines (iPSCs) and derive cardiomyocytes (iPSC-CMs). Aim 2a
utilizes a coimmunoprecipitation WB to assay disrupted SHROOM3-DVL2 binding within the variant iPSC-CM
lines. Aim 2b assays the variant iPSC-CMs for WB and IHC evidence of PCP disruption. Aim2c assays the
variant iPSC-CMs for altered PCP transcription using RNA-seq. The research plan will demonstrate SHROOM3's
mechanism as a novel component of the PCP pathway and a novel CHD candidate gene. Indiana University's
top10 NIH funded Department of Pediatrics provides a highly supportive environment for physician-scientist
development, and my unique panel of mentors each has world-class expertise in an important aspect of the
project, including cardiovascular genetics, development, and disease modeling with mice and iPSCs. I will gain
a skillset in statistics, molecular and developmental biology, mouse and iPSC disease models, CRISPR-Cas9
gene editing and genomics...

## Key facts

- **NIH application ID:** 10450640
- **Project number:** 5K08HL148508-03
- **Recipient organization:** INDIANA UNIVERSITY INDIANAPOLIS
- **Principal Investigator:** Matthew D. Durbin
- **Activity code:** K08 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $155,248
- **Award type:** 5
- **Project period:** 2020-07-07 → 2025-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10450640

## Citation

> US National Institutes of Health, RePORTER application 10450640, "SHROOM3 is a Novel Cause of Congenital Heart Disease" (5K08HL148508-03). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10450640. Licensed CC0.

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