MicroRNA Mirn140 modulates Pdgf signaling during palatogenesis

Abstract

Disruption of signaling pathways such as those mediated by Shh or Pdgf causes craniofacial disease, including cleft palate. The role that microRNAs play in modulating palatogenesis, however, is completely unknown. We show, in zebrafish, that the microRNA Mirn140 negatively regulates Pdgf signaling during palatal development and we provide a mechanism for how disruption of Pdgf signaling causes palatal clefting. The pdgf-receptor alpha (pdgfra) 3' UTR contains a Mirn140 binding site functioning in the negative regulation of Pdgfra protein levels in vivo. Both pdgfra mutants and Mirn140-injected embryos share panoply of facial defects including clefting of the crestderived cartilages that develop in the roof of the larval mouth. Concomitantly, the oral ectoderm beneath where these cartilages develop loses pitx2 and shha expression. Mirn140 modulates Pdgfmediated attraction of cranial neural crest cells to the oral ectoderm, where crest-derived signals are necessary for oral ectodermal gene expression. Both Mirn140 loss-of-function and pdgfra overexpression alters palatal shape and causes neural crest cells to accumulate around the optic stalk, a source of the ligand Pdgfaa. Conserved molecular genetics and expression patterns of mirn140 and pdgfra suggest that their regulatory interactions are ancient methods of palatogenesis that provide a candidate mechanism for cleft palate.Cleft palate and other craniofacial diseases are common in humans and have complex cellular and genetic etiologies. In amniotes, the palate serves to separate the nasal and oral cavities and is generated through an intricate series of morphogenic events that include early neural crest cell migration and cell-cell signaling during the formation of facial prominences, as well as later generation and fusion of palatal shelves. While later events involving palatal shelves have not been described in zebrafish, palatal precursors migrate both rostral and caudal to the eye to condense upon the oral ectoderm in amniotes 1 as well as zebrafish 2,3 and evidence continues to accumulate that the early signaling environment governing palatogenesis is also largely equivalent [3][4][5][6] . For instance, Hh signaling is crucial for palatogenesis in humans and zebrafish 3,4,7 . Zebrafish and amniotes also share expression patterns of palatogenic genes such as Shh 4,8 , Fgf8 4,9,10 and Pdgf receptor alpha (Pdgfra) [11][12][13] .In mouse, the Pdgf family consists of four soluble ligands, Pdgfa, Pdgfb, Pdgfc, and Pdgfd as well as two receptor tyrosine kinases, Pdgfra and Pdgfrb 14 . Pdgf signaling regulates a myriad of biological processes as demonstrated by analyses of mouse Pdgf ligand and receptor mutants 14 . Mice null for Pdgfra have a facial clefting phenotype that includes cleft palate 12,13 . This facial phenotype is fully recapitulated in mice doubly mutant for Pdgfa and Pdgfc 15. Most Pdgfc mutants have cleft palate 15 MicroRNAs (miRNAs) provide a unique mechanism for modulating signaling pathways [17][18][19][20] . Skeletogenic, including...

MicroRNAs as regulators of bone homeostasis and bone metastasis

“…MiR-140, a positive regulator of chondrogenesis through the inhibition of HDAC4 and Dnpep, contributes to craniofacial development and endochondral bone formation. [14][15][16] Consistent with this, mice lacking miR-140 display accelerated bone formation at embryonic and neonatal stages of development. 15 Conversely, miR-199a* and miR-145 have been shown to negatively regulate chondrocyte differentiation; miR-199a* downregulates the expression of Smad1, 17 whereas miR-145 targets Sox9, a key transcription factor involved in chondrogenesis.…”

PDGFRa mutations in humans with isolated cleft palate

“…Disruption of platelet-derived growth factor receptor alpha (Pdgfra) in zebrafish by Mirn140 was found to cause craniofacial abnormalities including cleft palate. 6 Consistent evidence supporting a significant role in palatal development came from previous studies in Pdgfra knockout mice. [7][8][9][10] PDGFRa (MIM 173490) maps to chromosome 4q12, spans 65 kb, contains 23 exons, and encodes a protein with 743 amino-acid residues.…”

“…Pdgfra mutations and Mirn 140-injected embryos shared a range of facial defects, including cleft palate. 6 A recent report suggests that the single-nucleotide polymorphisms (SNP) region in pre-miR-140 contributes to cleft palate susceptibility by influencing the processing of miR-140 in human PDGFRa. 17 In addition, a subsequent publication has revealed possible synergistic interaction between infants with CA/AA at rs7205289 located in the microRNA (miR)-140 and maternal passive smoking in contributing to cleft palate risk.…”

Bone marrow stroma-derived miRNAs as regulators, biomarkers and therapeutic targets of bone metastasis

“…MiR-140 positively regulates chondrocyte differentiation by inhibiting HDAC4 and Dnpep, and contributes to craniofacial development and endochondral bone formation. 17,18 Consistent with this, mice lacking miR-140 display accelerated bone formation at the embryonic and neonatal stages of development. 18 Conversely, miR-199a* and miR-145 negatively regulate chondrogenesis; miR-199a* downregulates the expression of Smad1, 19 whereas miR-145 targets Sox9, which encodes an essential transcription factor for chondrocyte differentiation.…”