Molecular Regulation of Palate Development

Abstract Submucous cleft palate (SMCP), defined by abnormal attachment of palatal muscles with intact oral and nasal mucosa, is a common craniofacial birth defect that disrupts the essential function of the soft palate for swallowing, causing feeding difficulties, middle ear dysfunction leading to hearing loss, and lifetime speech problems. Humans have five pairs of palatal muscles that all attach to the palatine aponeurosis, a fan-like dense fibrous connective tissue comprising the core of the soft palate that connects to the hard palate bone. Whereas the palatal muscles are derived from embryonic pharyngeal mesoderm, all connective tissues in the soft palate, including the palatine aponeurosis, tendons, and the muscle connective tissues that are embedded within and ensheathing the muscles, develop from cranial neural crest cell (CNCC) derived mesenchyme. Studies of limb muscle development have identified the embryonic muscle connective tissue as an important source of signals for patterning both the nascent muscles and tendons. However, how muscle connective tissue and tendon cells are specified from a common mesenchymal progenitor population and how they regulate muscle and tendon development remain unclear. We found that the Foxf2 and Foxd1 transcription factors exhibit complementary and partly overlapping patterns of expression along the medial- lateral axis of the embryonic soft palate mesenchyme at the onset of palatal myogenesis, with Foxf2 expression enriched in medial mesenchymal progenitors and with Foxd1 strongly expressed in the mesenchyme cells in the muscle-forming lateral region of the embryonic soft palate in mice. We demonstrate that Foxf2-deficient mice exhibit specific soft palate defects, including mispositioned palatal muscles and failure of palatine aponeurosis formation. Expression of Foxd1 and muscle-associating mesenchyme markers are ectopically activated in the medial mesenchymal progenitor cells in Foxf2 mutant embryonic soft palate. Furthermore, Foxd1 heterozygosity substantially rescues palatine aponeurosis formation and integration with the tensor veli palatini (TVP) muscles in Foxf2 mutant mice. On the other hand, homozygous disruption of Foxd1 causes aberrant splitting and abnormal attachment of TVP muscles to the pterygoid plate. Our central hypothesis is that Foxf2 and Foxd1 act antagonistically to regulate the specification and differentiation of distinct connective tissue lineages from the CNCC-derived soft palate mesenchyme to guide the morphogenesis and integration of palatal muscles. We propose comprehensive experimental studies to gain unprecedented understanding of the cellular and molecular mechanisms regulating soft palate connective tissue development and palatal muscle morphogenesis. Data from these studies provide a rich resource for uncovering mechanisms coordinating development and integration of musculoskeletal tissues and will lead to improvement in diagnosis and treatment of SMCP and other musculoskeletal disorders.