The Journal of Neuroscience, March 22, 2006, ():
Notch1 Signaling Regulates Radial Glia Differentiation through Multiple Transcriptional Mechanisms
J. Neurosci. Patten et al.
26: 3102
Supplemental data
Files in this Data Supplement:
- supplemental material
-
Supplementary Figure 1. Contactin does not regulate radial glia morphological differentiation or Notch1 transcriptional events in glia. Soluble contactin was purified from conditioned media of cells over expressing a rat Ig-contactin construct (see Peles et al., 1995). A, To test if the purified contactin was functional we examined if it upregulates MAG expression in an oligodendrocyte cell line (OLN-93), as previously shown (Hu et al., 2003). Cells were treated with contactin (1:500) for 48 hours, lysed and subjected to western blot analysis with an anti-MAG antibody. The blot was then stripped and probed with an anti-GAPDH antibody for normalization. Contactin treatment increased MAG expression indicating this soluble contactin was functional. B, C To determine the role of contactin in radial glia morphological differentiation, glia were either transfected with NICD (for a positive control), left untreated (control) or treated with contactin (1:500) for twenty-four hours. Images show representative glia under control conditions or treated with contactin demonstrating that contactin had no effect on radial glial morphology (B). Quantification of the morphology revealed no effect of contactin on inducing radial glia morphology. Asterisk marks which condition is significantly different from control, p < 0.03 (C). D, E, F, To test the role of contactin on activating Notch1 transcriptional mechanisms, purified P6 cerebellar glia were co-transfected with plasmids encoding the erbB2 (D), CBF1 (E) or BLBP (F) luciferase reporters and a control plasmid. NICD was used as a positive control. Cells were then treated with contactin at the indicated concentrations and twenty-four hours later the cells were lysed and luciferase activity measured. Contactin treatment had no effect on Notch1 transcriptional events. Asterisks mark which conditions are significantly different from control, p < 0.04. AU, Arbitrary units.
