# Genes and cell populations involved in orofacial clefting

> **NIH NIH R03** · UNIVERSITY OF COLORADO DENVER · 2020 · $155,500

## Abstract

Project Summary
Fusion of the facial prominences to form the upper lip and primary palate is a critical part of normal human
development and failure of this process results in a type of orofacial clefting affecting ~1.5 in 1,000 human
births termed cleft lip with or without cleft palate (CL+/-P). Despite the importance of this process, we only
have limited knowledge of the genes and cell types that are responsible for carrying out successful fusion of
the facial prominences at a critical region termed the lambdoid junction, where the medial nasal process meets
with the lateral nasal process and maxillary prominence. Its small size and complex organization have made
this fusion zone difficult to isolate and study using conventional gene expression analyses. With single cell
RNA sequencing (scRNA-seq), we assessed the cell types, cell behaviors, and gene expression profiles for the
wild-type lambdoid junction. We identified particular cell types associated with fusion and specific gene
signatures expressed within these populations. Importantly, many of these genes have been relatively
understudied in model organisms. Building on these preliminary data, the goal of this proposal is to investigate
if and how these cell populations and their gene signatures are altered in mouse models of human clefting.
First, I will study if and how the expression of the marker genes of the cell populations at the fusion zone is
altered in a small set of mouse models of human orofacial clefting. I will use models that are: (a) of full
penetrance and invariable expressivity; (b) clearly involving the l junction in any pathology; (c) relevant to
human orofacial clefting; and (d) that represent different scenarios of l junction fusion failure. Specifically, I will
use Trp63, Bmpr1a, Tfap2a, and Aldh1a3 models of clefting and craniofacial dysmorphology. For each of
these models, I will assess the expression of the marker genes by both in situ hybridization and by qRT-PCR,
and assess if there are similarities and differences in how these genes and their associated cell populations
are altered compared to control embryos. Secondly, I will perform lambdoid junction scRNA-seq on one of the
models to obtain a more in-depth analysis of cell populations and gene expression. The region corresponding
to the lambdoid junction will be isolated from stage-matched wild-type and mutant embryos, and dissociated
into single cells for sequencing in a routine and reproducible procedure. Subsequently, standard bioinformatics
procedures will be employed to assess if and how the cell populations and gene expression profiles of the
relative cell populations have been altered in the cleft lip model. Finally, any differences will be verified and
extended to the other models using in situ hybridization and qRT-PCR to determine if similar or different
mechanisms underlie these various mouse clefting models. The work in this proposal will provide a crucial
understanding for how orofacial clefting ...

## Key facts

- **NIH application ID:** 9882263
- **Project number:** 5R03DE028635-02
- **Recipient organization:** UNIVERSITY OF COLORADO DENVER
- **Principal Investigator:** Hong Li
- **Activity code:** R03 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $155,500
- **Award type:** 5
- **Project period:** 2019-03-01 → 2021-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9882263

## Citation

> US National Institutes of Health, RePORTER application 9882263, Genes and cell populations involved in orofacial clefting (5R03DE028635-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9882263. Licensed CC0.

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