The Journal of Neuroscience, October 6, 2004, ():
Laminar Patterning in the Developing Neocortex by Temporally Coordinated Fibroblast Growth Factor Signaling
J. Neurosci. Hasegawa et al.
24: 8711
Supplemental data
Files in this Data Supplement:
- supplemental material
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Supplemental Figure 1. Transcriptional activation by Erm, Er81 and Pea3 and transcriptional repression by DN-Erm, DN-Er81, and DN-Pea3
Quantitative analyses of transcriptional activities of each Pea3-Ets transcription factor form used for the in vivo electroporation-mediated gene transfer were carried out by luciferase reporter assays to assess the cross-reactivity of each DN-form with other members of Pea3-Ets transcription factors. Among Pea3-Ets transcription factors, Erm increased luciferase activity from the reporter plasmid to levels 2 to 3 fold higher than that achieved by reporter plasmid alone. In contrast, Er81 and Pea3 only increased the activity to levels 1.5 to 2.2 fold higher. Nevertheless, each DN-form decreased transcriptional activities achieved by the full-length Pea3-Ets transcription factors to levels achieved by reporter plasmid alone. Furthermore, they cross-reacted with all the other Pea3-Ets transcription factors and suppressed their transcriptional activities to a similar extent, as expected from their sequence similarity (amino acids with 95% sequence identity). Comparative luciferase activity is shown as a percentage (mean ± sem) normalized to the luciferase activities achieved by the expression of Pea3-Ets transcription factor responsive reporter alone. Each data point was measured and calculated from >5 independent transfections
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Supplemental Figure 2. Immunohistochemical analyses of cortical descendants that are stalled in the intermediate zone after the electroporation
Immunohistochemical analyses of DN-Erm incorporated (A) and FGFR3c-?C incorporated (B) cortical descendants that were accumulated in the intermediate zone. EYFP positive descendants (green in color) were positive for both ?III-tubulin (red in color) and MAP2 (red in color) expressions. Expression of DN-Er81 and DN-Pea3 also gave cortical descendants that are positive with ?III-tubulin and MAP2 (data not shown). Scale bar: 10 ?m.
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Supplemental Figure 3. In situ hybridization analyses of FGFRs expressed in the neocortical VZ
Coronal sections of E12.5 (A, D, G), E14.5 (B, E, H) and E16.5 (C, F, I) cerebral cortices, showing expression patterns of FGFR1c (A-C), FGFR2c (D-F), and FGFR3c (G-I), determined by in situ hybridization. Scale bar; 0.16 mm for (A, D, G): 0.4 mm for (B, C, E, F, H, I).
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Supplemental Figure 4. Summary diagram
Diagram indicating a feedback mechanism of how the early-late temporal sequence of cortical neuron generation in the VZ is linked with the deep-superficial spatial sequence of cortical neuron positioning in the CP. PE: early-progenitors; PM: mid-progenitors; PL: late-progenitors; ND: deep-layer cortical neurons; NI: intermediate-layer cortical neurons; NS: superficial-layer cortical neurons.
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