REST Regulates Distinct Transcriptional Networks in Embryonic and Neural Stem Cells
Figure 1
Validation of Stem Cell Potency and Microarray Sensitivity
(A) Mouse ESC stain for markers of self-renewal. Immunohistochemistry was carried out using antibodies to Oct4, Nanog, and Sox2. For negative control (-ve), the primary antibody was omitted.
(B and C) ESC are capable of differentiation. Following treatment with retinoic acid (RA), qRT-PCR was used to assay expression of self-renewal markers (Nanog, Oct4) (B) and differentiation markers (Alpha fetoprotein, Afp; Coup TFII, Nr2f2).
(D) NSC are multipotent. Following a standard differentiation protocol (see Text S1), cells were observed to differentiate into neurons, astrocytes and oligodendrocytes, as revealed by staining with Tuj1 (βIII tubulin), GFAP (glial fibrillary acidic protein) and O4 antibodies, respectively. Scale bar: 20 μm.
(E) Enrichment for REST occupancy in ESC at the Syt4 RE1 site as determined by ChIP-chip. Locations of seven tiled probes and RE1 site are indicated.
(F) Comparative performance of ChIP-chip and qPCR for 150 randomly selected RE1s. Dark blue (occupied) and gray bars (unoccupied) indicate a consistent call by both techniques. Light blue bars denote RE1s that were not consistently called.