Xenbase Image ID: 83476

Figure 5. HIRA Downregulation Resembles the H3.3 Phenotype (A) Specificity of HIRA morpholinos. (Left) The scheme shows injections of HIRA MO (92 ng) in fertilized eggs. After incubation to reach stage 11 in controls, 40 noninjected () and 40 injected (MO HIRA) embryos were used to prepare soluble extracts for western blot analysis. (Right) Results used a 2-fold dilution series (gradient bar) with the highest quantity being equivalent to three embryos, and detection with a HIRA antibody. Anti-β-actin and memcode staining served as loading controls. (B) Downregulation of HIRA but not WDR5 leads to gastrulation defects. We injected the indicated MO (46 ng) in one cell of 2-cell stage embryos. We acquired images when controls were at the indicated stages. White arrows indicate the gastrulation defects observed in HIRA morphants. In the bottom panel, we show a broader coverage of the gastrulation time period (stages 11 to 13) with eight distinct time points (see also Movie S1). Scale bar: 1 mm. (C) Xbra mRNA in situ hybridization in HIRA- and WDR5-deficient embryos. We injected the indicated MO (69 ng for CTL or WDR5, 46 ng for either HIRA or mutHIRA) in one cell of 4-cell stage embryos. After incubation to reach the indicated stages in controls, we fixed injected embryos and performed whole-mount in situ hybridization with an Xbra probe. White arrows indicate the injected part and arrowheads the noninjected part of HIRA morphant embryos. Scale bar: 1 mm. See also Figures S3 and S5. Image published in: Szenker E et al. (2012) Copyright © 2012. Image reproduced with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License. Gene Synonyms Species Stage(s) Tissue tbxt.S bra, brachyury, ntl, t, t-a, t-b, X-bra, Xbra, Xbrachyury X. laevis Sometime during NF stage 10.5 to NF stage 23 mesoderm Image source: Published Experiment + Assay Source Phenotypes and Disease Xla Wt + hira MO + NF11-24 (morphology) fig.5.b Anatomical Phenotype abnormal gastrulation abnormally delayed closure of blastopore Xla Wt + hira MO + NF24-28 (immunohistochemistry) fig.5.b Anatomical Phenotype increased apoptosis in embryo Xla Wt + hira MO + NF24-28 (morphology) fig.5.b Anatomical Phenotype decreased length of anterior-posterior axis Xla Wt + wdr5 MO + NF24-28 (morphology) fig.5.b Anatomical Phenotype decreased length of anterior-posterior axis decreased size of the head decreased size of the tail Xla Wt + hira MO + NF10.5 (in situ hybridization) fig.5.c Expression Phenotype decreased amount tbxt.S expression in mesoderm Larger Image Printer Friendly View

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