Abstract Platelet-Derived Growth Factor Receptor β (PDGFRβ) is a receptor tyrosine kinase crucial for the survival, proliferation, differentiation and migration of mesenchymal cells in development. PDGF binding to PDGFRβ induces receptor dimerization, which allows a conformational change in the kinase domain of the receptor and autophosphorylation of tyrosine residues on the PDGFRβ intracellular domain. Activating mutations in PDGFRB have been recently discovered in humans. These activating mutations cause the receptor kinase domain to remain in a constitutively active state even without ligand binding or receptor dimerization. Penttinen Syndrome is an ultra-rare disease caused by a V665A point mutation in the kinase domain of PDGFRβ. Craniofacial phenotypes appear during childhood including craniosynostosis, thin calvarium, Wormian bones, and midface retrusion. Little is currently known regarding the full range of phenotypes associated with the V665A mutation. Even less is known about how this excess of PDGFRβ signaling leads to human disease. To address this gap in knowledge, a mouse model with a conditional V665A mutation in the Pdgfrb gene has been generated. Reminiscent of Penttinen Syndrome patients, V665A mice (V) exhibit craniofacial defects. Additionally, preliminary studies show that signal transducer and activator of transcription 1 (STAT1) is overexpressed and phosphorylated in cranial mesenchyme. It is hypothesized that constitutive PDGFRβ-V665A signaling through STAT1 interferes with cranial development by inhibiting key transcription factors required for mesenchymal progenitors to form meninges. First, this proposal will identify the cellular mechanisms of craniofacial defects in V665A mutants (Aim 1) by assessing apoptosis, proliferation, differentiation, and migration of cranial mesenchyme during embryo development, by investigating whether these processes are normalized by deletion of STAT1, and using lineage-specific Cre mice to determine the cellular origin of the defects. Second, it will identify molecular mechanisms (Aim 2) using bulk RNA sequencing and Cleavage Under Targets & Release Using Nuclease (CUT&RUN) on cranial mesenchyme from control and PDGFR?-V665A mutant embryos. These studies will expand the current understanding of PDGFRβ signaling and will illuminate how elevated signaling leads to craniofacial defects in humans. Furthermore, the results of these studies may inform the adjacent fields of cancer and cardiovascular disease as there is evidence of upregulated PDGF signaling in these diseases.