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Figure 4

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ZDB-FIG-231205-4

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Wilcockson et al., 2023 - An improved Erk biosensor detects oscillatory Erk dynamics driven by mitotic erasure during early development

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Figure 4

modErk-KTR specifically reports on Fgf/Erk activity in early zebrafish embryos

(A and B) Stills of live ubiP:Erk-KTR-Clover transgenic embryos. Embryos are false-colored to indicate Erk activity levels; (green, high activity; magenta, low activity). Embryos are shown from an animal (A) or lateral (B) view. Insets show a magnified view of the region within the black box without false coloring. White-dashed line, embryo proper.

(Ai) Single z-slices through the center of embryos in (A) showing Erk activity around the embryonic margin using the same color scheme as (A).

(C) Live imaging of ubiP:Erk-KTR-Clover transgenic embryos following treatment with DMSO (control), 10 μM PD-0325901 (MEKi), 20 μM RO-3306 (CDK1i), or both MEKi and CDK1i for an hour from 4.0 hpf.

(D) Quantification of Erk activity in (C) at the margin and animally. Shown are the single-cell readouts of Erk activity overlayed with the per embryo averages and the overall mean for DMSO (control, n = 209 cells from 5 embryos [margin] or n = 200 cells from 5 embryos [animal]), 10 μM PD-0325901 (MEKi; n = 137 cells from 4 embryos [margin] or n = 196 from 5 embryos [animal]), 20 μM RO-3306 (CDK1i; n = 227 cells from 6 embryos [margin] or n = 133 cells from 4 embryos [animal]) or both 10 μM PD-0325901 and 20 μM RO-3306 (MEKi + CDK1i; n = 183 cells from 5 embryos [margin] or n = 194 cells from 5 embryos [animal]) treated embryos.

(E) Live imaging as in (C) of embryos injected with either 25 pg fgf8a or 500 pg dnFGFR at one-cell stage. Embryos were imaged at 50% epiboly (5.3 hpf).

(F) Quantification of Erk activity in (E) as in (D) for control (n = 142 cells from 3 embryos [margin] and n = 116 cells from 3 embryos [animal]), fgf8a (n = 130 cells from 4 embryos [margin], or n = 192 cells from 4 embryos [animal]) or dnFGFR (n = 168 cells from 4 embryos [margin] or n = 157 cells from 4 embryos [animal]) injected embryos.

Statistical tests were one-way ANOVA with Šidák’s multiple comparisons test.

Scale bars, 100 μm (A and B), 50 μm (Ai) or 20 μm (C–E); **** p > 0.0001.

See also Figure S1.

Expression Data

Expression Detail

Antibody Labeling

Phenotype Data

Phenotype Detail

Acknowledgments

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Reprinted from Developmental Cell, 58(23), Wilcockson, S.G., Guglielmi, L., Araguas Rodriguez, P., Amoyel, M., Hill, C.S., An improved Erk biosensor detects oscillatory Erk dynamics driven by mitotic erasure during early development, 2802-2818.e5, Copyright (2023) with permission from Elsevier. Full text @ Dev. Cell