Blocking zygotic transcription impairs heterochromatin. a Western blot for H3K9me3 (top) and α-tubulin (bottom) using protein extracted from embryos that were either mock injected (−) or injected with 0.2 ng of α-amanitin (+) at the one-cell stage. Protein was collected for analysis at 4.5 h post fertilization (hpf). b Chromatin immunoprecipitation with sequencing (ChIP) for H3K9me3 enrichment at pericentromeric Satellite-1 (Sat1) repeats in wild-type and α-amanatin-injected embryos. Embryos were collected for analysis at 4.5 hpf. P values were calculated using the Student’s t test; error bars indicate SEM. c Transmission electron microscopy (TEM) images demonstrating a lack of condensed chromatin ultrastructure in 5.0 hpf embryos that were injected with α-amanatin at the one-cell stage. Bottom panels represent higher magnification images (×20,000) of nuclear interior in mock- and α-amanatin-injected embryos at specified time points. Scale bars indicate 1 μm. d, e Quantification of the number of electron-dense aggregates per nuclear μm2 (d) and percent nuclear area covered by aggregates (e) in mock- and α-amanitin-injected embryos at 5.0 hpf. Particles/μm2 and percent nuclear area were measured in nuclei from each of four α-amanitin- and four mock-injected embryos. Each graphed data point represents data from one embryo; values for each embryo are the average of six to ten representative nuclei. Error bars indicate SD. Source data for panel a is provided as a source data file
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