vwa1 Knockout in Zebrafish Causes Abnormal Craniofacial Chondrogenesis by Regulating FGF Pathway
Author:
Niu Xiaomin1, Zhang Fuyu2, Ping Lu2ORCID, Wang Yibei3, Zhang Bo4, Wang Jian1, Chen Xiaowei1
Affiliation:
1. Department of Otolaryngology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China 2. 8-Year MD Program, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China 3. Department of Otolaryngology-Head & Neck Surgery, China-Japan Friendship Hospital, Beijing 100730, China 4. Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing 100730, China
Abstract
Hemifacial microsomia (HFM), a rare disorder of first- and second-pharyngeal arch development, has been linked to a point mutation in VWA1 (von Willebrand factor A domain containing 1), encoding the protein WARP in a five-generation pedigree. However, how the VWA1 mutation relates to the pathogenesis of HFM is largely unknown. Here, we sought to elucidate the effects of the VWA1 mutation at the molecular level by generating a vwa1-knockout zebrafish line using CRISPR/Cas9. Mutants and crispants showed cartilage dysmorphologies, including hypoplastic Meckel’s cartilage and palatoquadrate cartilage, malformed ceratohyal with widened angle, and deformed or absent ceratobranchial cartilages. Chondrocytes exhibited a smaller size and aspect ratio and were aligned irregularly. In situ hybridization and RT-qPCR showed a decrease in barx1 and col2a1a expression, indicating abnormal cranial neural crest cell (CNCC) condensation and differentiation. CNCC proliferation and survival were also impaired in the mutants. Expression of FGF pathway components, including fgf8a, fgfr1, fgfr2, fgfr3, fgfr4, and runx2a, was decreased, implying a role for VWA1 in regulating FGF signaling. Our results demonstrate that VWA1 is essential for zebrafish chondrogenesis through effects on condensation, differentiation, proliferation, and apoptosis of CNCCs, and likely impacts chondrogenesis through regulation of the FGF pathway.
Funder
National Natural Science Foundation of China Youth Programs of the National Natural Science Foundation of China
Subject
Genetics (clinical),Genetics
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