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Results: 4

1.
Figure 4

Figure 4. From: Multiple Signaling Pathways Converge on the Cbfa1/Runx2 Transcription Factor to Regulate Osteoblast Differentiation.

Overview of signal transduction pathways affecting Runx2 activity. Refer to text for explanation.

Renny T. Franceschi, et al. Connect Tissue Res. ;44(Suppl 1):109-116.
2.
Figure 3

Figure 3. From: Multiple Signaling Pathways Converge on the Cbfa1/Runx2 Transcription Factor to Regulate Osteoblast Differentiation.

Synergistic stimulation of osteoblast gene expression by transduction of mesenchymal cells with adenoviruses overexpressing Runx2 and BMPs. (A) Alkaline phosphatase activity. C3H10T1/2 cells were transduced with optimal titers of adenoviruses containing a LacZ control cDNA (LacZ) or cDNAs encoding BMPs 2, 4, or 7 and/or Runx2 (R) as indicated. Cells were harvested after 6 days and assayed for alkaline phosphatase activity. (B) mRNA levels. Cells were transduced with AdLacZ, AdBmp2, or AdRunx2 adenovirus as indicated. After various times, total RNA was isolated for Northern blot analysis of Runx2, and OCN mRNAs as well as for 18S rRNA for blot normalization.

Renny T. Franceschi, et al. Connect Tissue Res. ;44(Suppl 1):109-116.
3.
Figure 2

Figure 2. From: Multiple Signaling Pathways Converge on the Cbfa1/Runx2 Transcription Factor to Regulate Osteoblast Differentiation.

FGF2-dependent activation and phosphorylation of Runx2. (A) Activation of the OCN promoter. MC3T3-E1 preosteoblast cells stably transfected with a 1.3 kb fragment of the mouse osteocalcin gene 2 promoter driving a firefly luciferase reporter gene were treated with vehicle or 12.5 ng/mL FGF2 for 6 hr in the presence or absence U0126, a MEK inhibitor, or the inactive analogue, U0124. (B) Runx2 phosphorylation. MC3T3-E1 cells were metabolically labeled with 32P orthophosphate and treated with the indicated concentrations of FGF2 for 6 hr in the presence or absence of U0126 or U0124. Endogenous Runx2 was immunoprecipitated with a polyclonal antibody and analyzed by SDS-PAGE and autoradiography (upper panel). Total Runx2 in immunoprecipitates was measured by Western blotting (lower panel).

Renny T. Franceschi, et al. Connect Tissue Res. ;44(Suppl 1):109-116.
4.
Figure 1

Figure 1. From: Multiple Signaling Pathways Converge on the Cbfa1/Runx2 Transcription Factor to Regulate Osteoblast Differentiation.

Activation of the MAPK pathway stimulates phosphorylation within the P/S/T domain of Runx2. (A) Runx2 domains. Diagram shows the three major domains of Runx2: the glutamine/alanine-rich region (Q/A), the Runt domain that contains DNA binding and nuclear localization sequences (NLS), and the proline/serine/threonine (P/S/T) domains. Inset shows the amino acid sequence of a region of the P/S/T domain shown to be crucial for activation of transcription by the MAPK pathway. Consensus MAPK phosphorylation sites are indicated by asterisks. (B, C) Runx2 phosphorylation. Wild-type Runx2 (Panel B) or a P/S/T domain deletion mutant (ΔPST, Panel C) both containing a N-terminal HA tag were expressed in COS7 cells using a pcDNA5 expression vector. Cells were co-transfected with either pcDNA5 containing a β-galactosidase cDNA (β-gal) or a constitutively active MEK cDNA, MEK(SP). Cells were metabolically labeled with either 32P orthophosphate or 35S methionine/cysteine, and cell extracts immunoprecipitated with an anti-HA epitope monoclonal antibody. Immunoprecipitates were analyzed by SDS-PAGE and autoradiography.

Renny T. Franceschi, et al. Connect Tissue Res. ;44(Suppl 1):109-116.

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