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. 2010 Apr 5;189(1):159–170. doi: 10.1083/jcb.200908048

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© 2010 Kohyama et al.

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Figure 4. — Astrocytes express functional REST/NRSF. (A) Expression of REST/NRSF in GFAP-positive astrocytes derived from P0 mouse brain. Bar, 50 µm. Inset: Hoechst nuclear staining of this field. (B) Expression of REST/NRSF in astrocytes in vivo. REST/NRSF (green) was expressed in GS (red)-positive cells in the cortical surface region of P8 mouse brain. Bar, 50 µm. Inset: Hoechst nuclear staining of the field. (C) REST/NRSF activity was examined using a reporter construct carrying the SCG10 minimal promoter (pGL3-S10PR) with intact (S36+) and mutated (Sm36+) RE1/NRSE. S36+ reporter activity was derepressed by the expression of DN-REST/NRSF in astrocytes (left). The reporter activity was enhanced by mutating the RE1/NRSE motif (middle). Mean ± SD (n = 3). Statistical significance was examined by Student’s t test (*, P < 0.05). Endogenous expression of SCG10 was also examined by RT-PCR analysis in astrocytes introduced with control or DN-REST/NRSF-expressing vector (right). (D) Transcriptional suppressor activity of REST/NRSF was observed in astrocytes and ES cells as assessed by the RE1/NRSE reporter system. Mean ± SD (n = 3). Statistical significance was examined by Student’s t test (*, P < 0.05). (E) Comparison, between neurons and astrocytes, of REST/NRSF association with RE1/NRSE-containing regions in GAD1, BDNF, Synaptotagmin4 (Syt4), and Calbindin (Cal) genes, measured by quantitative ChIP analysis. Blue columns: astrocytes. Red columns: neurons. Mean ± SD.