Nell-1, a key functional mediator of Runx2, partially rescues calvarial defects in Runx2(+/-) mice

J Bone Miner Res. 2011 Apr;26(4):777-91. doi: 10.1002/jbmr.267.

Abstract

Mesenchymal stem cell commitment to an osteoprogenitor lineage requires the activity of Runx2, a molecule implicated in the etiopathology of multiple congenital craniofacial anomalies. Through promoter analyses, we have recently identified a new direct transcriptional target of Runx2, Nell-1, a craniosynostosis (CS)-associated molecule with potent osteogenic properties. This study investigated the mechanistic and functional relationship between Nell-1 and Runx2 in regulating osteoblast differentiation. The results showed that spatiotemporal distribution and expression levels of Nell-1 correlated closely with those of endogenous Runx2 during craniofacial development. Phenotypically, cross-mating Nell-1 overexpression transgenic (CMV-Nell-1) mice with Runx2 haploinsufficient (Runx2(+/-)) mice partially rescued the calvarial defects in the cleidocranial dysplasia (CCD)-like phenotype of Runx2(+/-) mice, whereas Nell-1 protein induced mineralization and bone formation in Runx2(+/-) but not Runx2(-/-) calvarial explants. Runx2-mediated osteoblastic gene expression and/or mineralization was severely reduced by Nell-1 siRNA oligos transfection into Runx2(+/+) newborn mouse calvarial cells (NMCCs) or in N-ethyl-N-nitrosourea (ENU)-induced Nell-1(-/-) NMCCs. Meanwhile, Nell-1 overexpression partially rescued osteoblastic gene expression but not mineralization in Runx2 null (Runx2(-/-)) NMCCs. Mechanistically, irrespective of Runx2 genotype, Nell-1 signaling activates ERK1/2 and JNK1 mitogen-activated protein kinase (MAPK) pathways in NMCCs and enhances Runx2 phosphorylation and activity when Runx2 is present. Collectively, these data demonstrate that Nell-1 is a critical downstream Runx2 functional mediator insofar as Runx2-regulated Nell-1 promotes osteoblastic differentiation through, in part, activation of MAPK and enhanced phosphorylation of Runx2, and Runx2 activity is significantly reduced when Nell-1 is blocked or absent.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkaline Phosphatase / genetics
  • Alkaline Phosphatase / metabolism
  • Animals
  • Animals, Newborn
  • Calcification, Physiologic / drug effects
  • Calcification, Physiologic / physiology
  • Calcium-Binding Proteins / genetics
  • Calcium-Binding Proteins / metabolism*
  • Calcium-Binding Proteins / pharmacology
  • Cell Differentiation / physiology
  • Cells, Cultured
  • Cleidocranial Dysplasia / genetics
  • Cleidocranial Dysplasia / metabolism
  • Cleidocranial Dysplasia / pathology
  • Core Binding Factor Alpha 1 Subunit / deficiency
  • Core Binding Factor Alpha 1 Subunit / genetics
  • Core Binding Factor Alpha 1 Subunit / metabolism*
  • Craniofacial Abnormalities / genetics*
  • Craniofacial Abnormalities / metabolism
  • Craniofacial Abnormalities / pathology
  • Dura Mater / metabolism
  • Extracellular Signal-Regulated MAP Kinases / antagonists & inhibitors
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • Femur / metabolism
  • Gene Expression / genetics
  • Glycoproteins / genetics
  • Glycoproteins / metabolism*
  • Glycoproteins / pharmacology
  • Haploinsufficiency / genetics
  • Heterozygote
  • JNK Mitogen-Activated Protein Kinases / antagonists & inhibitors
  • JNK Mitogen-Activated Protein Kinases / metabolism
  • Mice
  • Mice, Inbred Strains
  • Mice, Knockout
  • Mice, Transgenic
  • Osteoblasts / cytology
  • Osteoblasts / metabolism
  • Osteocalcin / genetics
  • Osteocalcin / metabolism
  • Osteogenesis / drug effects
  • Osteogenesis / physiology
  • Osteopontin / genetics
  • Osteopontin / metabolism
  • Phosphorylation / drug effects
  • Protein Kinase Inhibitors / pharmacology
  • RNA, Small Interfering / genetics
  • Skull / abnormalities*
  • Skull / cytology
  • Skull / embryology
  • Skull / metabolism
  • Skull / pathology
  • Sp7 Transcription Factor
  • Tibia / metabolism
  • Tissue Culture Techniques
  • Transcription Factors / genetics
  • p38 Mitogen-Activated Protein Kinases / antagonists & inhibitors
  • p38 Mitogen-Activated Protein Kinases / metabolism

Substances

  • Calcium-Binding Proteins
  • Core Binding Factor Alpha 1 Subunit
  • Glycoproteins
  • Nell1 protein, mouse
  • Protein Kinase Inhibitors
  • RNA, Small Interfering
  • Runx2 protein, mouse
  • Sp7 Transcription Factor
  • Sp7 protein, mouse
  • Transcription Factors
  • Osteocalcin
  • Osteopontin
  • Extracellular Signal-Regulated MAP Kinases
  • JNK Mitogen-Activated Protein Kinases
  • p38 Mitogen-Activated Protein Kinases
  • Alkaline Phosphatase