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Fig. 6. ß-catenin/TCF complex directly regulates the Ngn1 promoter.
(A) A schematic representation of the mouse neurogenin 1 (Ngn1)
promoter (WT), and its mutant within the putative TCF binding site (-1167 to
-1160) located in the Ngn1 promoter (Mut). NPCs were transfected with
a vector containing the Ngn1 promoter (2.7 kb: wild type or mutant)
driving luciferase expression. (B) Relative luciferase activity was measured
after 13 hours of incubation. Mutation of the TCF binding site in the
Ngn1 promoter reduced endogenous transcriptional activity. (C)
Chromatin complex was immunoprecipitated with anti-ß-catenin (Santa Cruz
Biotechnology and Sigma) or control IgG, and was subjected to PCR analysis to
amplify Ngn1 genomic sequence. (D) RT-PCR analysis of Ngn1
expression. NPCs were infected with a retrovirus encoding either GFP alone
(control) or both GFP and S33Y ß-catenin, and the expression level of
Ngn1 mRNA was analyzed. Gapdh was used for standardization
of the samples. No genomic amplification was observed from the RNA treated
without reverse transcriptase (-RT).
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