Cleft lip represents one of most common birth defects and affects approximately 1 in every 700 newborns worldwide. It is caused by disruption of normal upper lip development with environmental or genetic factors. The long term goal of this proposed research is to understand the mechanisms of upper lip development and of orofacial cleft pathogenesis. Mutations of Platelet Derived Growth Factor Receptor ? (PDGFR?) signaling have been tightly linked to cleft lip/palate in humans and mice, suggesting an evolutionarily conserved role in craniofacial development. During embryo development, the upper lip is formed by highly coordinated development of medial nasal process (MNP), lateral nasal process (LNP) and maxillary processes (MxP), all of which are originating from neural crest cells (NCCs). My previous study reveals that PDGFR? signaling is required to maintain MNP cell proliferation and regulate NCC migration, and indicates a role for small GTPase Rac1 in these processes downstream of PDGFR?. This proposed research is aimed at characterizing the role of Rac1 signaling in mediating PDGFR? regulation on MNP and NCC development.
In aim one, I will use a newly generated unbiased Wnt1-Cre2 allele, to ablate Rac1 function specifically in NCCs, and characterize the mutant craniofacial phenotype at histological, cellular and molecular levels. The epistatic effect between Rac1 and PDGFR? will be tested by inactivating a single allele of each gene in NCCs. I will further rescue the craniofacial phenotype of PDGFR? fl/fl;Wnt1-Cre2 mice by driving expression a constitutively active form of Rac1 in NCCs in vivo.
In aim two, I will examine how PDGFR? signaling regulates Rac1 activity during MNP morphogenesis. I will analyze the activity of immediate downstream signaling pathways in PDGFR? deficient MNP, and examine of the interaction between the identified signaling pathways and Rac1 activity.
Aim three is designed to characterize the expression of PDGFR? transcriptional targets Cdc42ep3 (CEP3) and its role in craniofacial development. CEP3 will be examined with in vitro assays and gene-targeting method. Results of the proposed works will delineate the signaling cascade by which PDGFR? regulates NCC and MNP development, at the levels of signaling transduction and transcription. The results of proposed research will provide novel information to understand the fundamental mechanisms of MNP development and upper lip morphogenesis. This study will hold important benefit in treatment of cleft lip, to ultimately reduce the occurrence of this birth defect in humans.
Cleft lip occurs once in every 700 new births worldwide, consisting one of most common birth defects with poorly understood mechanisms. Mutations of Platelet Derived Growth Factor (PDGF) signaling have been associated with orofacial clefting pathogenesis accompanying downregulation of Rac1 activity. The application has some minor weaknesses;however, this application is well-designed to delineate the mechanisms how Rac1 signaling mediates PDGFRa regulation on neural crest cell development during upper lip morphogenesis, and the results will provide novel information to understand the fundamental mechanisms of upper lip formation, and will benefit treatment and prevention of cleft lip in the future.