Magnetron Development

Contains reports on two research projects

Magnetron Development

Contains reports on three research projects

Microwave and Physical Electronics

Contains reports on six research projects

Microwave and Physical Electronics

Contains reports on seven research projects

PPARβ is essential for <i>X. laevis</i> development.

<p>(A) Design of the morpholino (PPARβ MO) to target PPARβ translation and of the control morpholino (Co). Capital letters designate nucleotides that can hybridize with the PPARβ MO. (B) Immunoblot showing endogenous PPARβ levels in non-injected embryos (Ni) and embryos injected with PPARβ MO or Co. β-actin served as a loading control. (C) Scoring of A–P axis defects. Different doses of PPARβ MO, Co, or a combination of PPARβ MO and PPARβ_rescue mRNA were injected. Embryos with a length about a third of that of non-injected sibling embryos were scored as ‘very-short axis’, and those with a length of about two thirds of normal were scored as ‘short axis’. (D) Representative not-injected (Ni), Co-injected (Co), MO-injected (MO), and MO combined with rescue injected (PPARβ MO + PPARβ_rescue) embryos.</p

PPARβ interprets a chromatin signature that is deposited at the end of the pluripotent stage.

<p>(A) Seven ‘K27’ genes and eight ‘K4 only’ genes were analysed by ChIP as indicated. Results are presented in a heat map (see also Supplementary Fig. 7). Variation in RNA expression upon PPARβ MO injection was obtained from the RNA-seq data or from qPCR validations. (B) ChIP with H3K27me3 antibody was conducted at stage 9 on 37 ‘PPARβ promoted genes’ and on 27 Control genes. PPARβ promoted genes were chosen among the top 200 most downregulated genes at stage 11, upon MO injection in the list presented in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083300#pone.0083300.s008" target="_blank">Table S1</a>, while Control genes did not show a change of expression upon MO injection. Results are presented as percentage of input. The threshold of 1% is indicated. Genes scored as positive for H3K27me3 are indicated by a red dot (see methods for further details on the definition of gene sets and on the criteria of scoring). (C) Sequential ChIPs were conducted. Note that no enrichment was observed for klf11 and for plcg1, which represent negative controls (see panel b). Error is the S.E.M of 2 independent experiments. (D) ChIP using PPARβ antibody was conducted at stage 11.5. Error is the S.E.M of 3 to 4 independent experiments. (E) ChIP with H3K27me3 antibody or PPARβ antibody and <i>q</i>RT-PCR were conducted on embryos treated with DZNep or DMSO and injected with PPARβ MO or Co. Error is the S.E.M of technical replicates of a single experiment that we have replicated with similar results.</p

Rate of transcript level variation is maximal at gastrula stage.

<p>(A) Data from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083300#pone.0083300-Irie1" target="_blank">[23]</a> were used to quantify transcription variations during normal development. The number of genes showing an RNA level increase or decrease by 2×, 4×, or 8× between two consecutive stages was plotted. Data were normalized by the duration, in hours, of each developmental period analysed. (B) The group of genes with RNA levels that increased 4× or more between stage 11 and stage 13 was considered, and the RNA levels of these genes were plotted at different developmental stages. The rectangles delineate the 25^th and 75^th percentiles, the horizontal bar is the median, and the whiskers indicate the 10^th and 90^th percentiles.</p

PPARβ promotes differentiation but represses dorsal mesoderm and endoderm specification.

<p>(A)–(C) Embryos were injected with PPARβ MO or Co, allowed to develop until stage 18 (A) or stage 11.5 (B) and (D), and collected for extraction of total RNA. qRT-PCR runs for a selection of neural (blue), mesodermal (red), or endodermal (yellow) markers of differentiation (A) and (B) or of germ layer specification (C) were conducted. RNA levels were normalized to EEF1a and RPL8 and are presented as fold variation between MO and Co samples. Error bars represent the S.E.M. of 3 to 5 independent experiments. (D) Embryos were injected with PPARβ MO or Co, fixed at stg. 11.5, hemi-sectioned along the dorso–ventral axis, and processed for RNA <i>in situ</i> hybridization. While Mo injection did not affect the <i>sox17α</i> expression domain, it resulted in the expansion of <i>brachyury</i> expression dorsally (see the scale) but not ventrally. Arrows indicate the dorsal lip. (E) Quantification of the surface covered by the dorsal and ventral expression domains of <i>brachyury</i> in MO compared to Co hemi-sections. Error bar is the S.E.M. of 10 measurements. *: two-tailed Student’s t-test vs control, P<0.05.</p