FIGURE

Fig. 5

ID
ZDB-FIG-191031-8
Publication
Prummel et al., 2019 - A conserved regulatory program initiates lateral plate mesoderm emergence across chordates
Other Figures
All Figure Page
Back to All Figure Page
Fig. 5

EomesA, FoxH1, and MixL1 together can induce the + 2.0drl enhancer.  a ChIP-seq tracks for EomesA. FoxH1, MixL1, and Smad2 in the  drl locus. See text for details. Bottom depicts the + 2.0drl intronic enhancer and the smaller, minimally specific region + 2.4drl after removal of repeat sequences.  b e Constitutively-active VP16-EomesA boosts + 2.0drl:EGFPreporter expression in its native territory. Compared to injection controls ( b c), microinjection of  VP16-eomesA mRNA in + 2.0drl: EGFP reporter transgenics enhances and prolongs EGFP expression in the native reporter expression domain ( d e).  f k Gastrulation-stage (shield to 75% epiboly) zebrafish embryos, lateral view left and animal view right, probed with  EGFP ISH to detect expression of + 2.0drl:EGFP. Compared to controls ( n = 18/18) ( f), embryos injected with mRNA encoding full-length  eomesA ( n = 16/26),  foxh1 ( n = 18/31), or  mixl1 ( n = 13/27) show enhanced + 2.0drl:EGFP reporter activity ( g i), as indicated by arrowheads. The asterisk in  g points out ectopic expression in the enveloping layer cells.  jCombining mRNAs encoding full-length  eomesA (e) foxh1 (f), and  mixl1 (m) ( e/f/m) triggers ectopic reporter expression also in dorsal blastomeres ( n = 13/15, compared to native reporter expression pattern in  f), an activity that also remained in embryos devoid of endoderm after  sox32 morpholino injection ( n = 7/17) ( k).  l p Mixl1 acts on the + 2.0drlenhancer as analyzed in Cas9 RNP-mediated crispants.  l Schematic representation of the  mixl1 and  mezzo loci, with the individual sgRNAs for mutagenesis annotated ( cc for  CRISPR cutting, followed by sgRNA index).  m p mRNA in situ hybridization of  EGFP expression in + 2.0drl: EGFP embryos as crispant control ( n = 30/30) ( m), injected with Cas9 RNPs of ( n mixl1 ccB ( n = 6/6), ( o mezzo ccA ( n = 11/11), and ( p mixl1 ccB together with  mezzo ccA sgRNA ( n = 21/27); the resulting mosaic  mixl1 ccB crispants show diminished + 2.0drl reporter expression at late gastrulation, as pointed out by arrowheads. Lateral and animal views as indicated. Scale bar in ( b) 500 μm, ( c) 250 μm


Expression Data

Expression Detail
Antibody Labeling
Phenotype Data

Phenotype Detail
Acknowledgments
This image is the copyrighted work of the attributed author or publisher, and ZFIN has permission only to display this image to its users. Additional permissions should be obtained from the applicable author or publisher of the image. Full text @ Nat. Commun.