Figure 4. Expression of selected presumptive direct targets of Six1/Eya1 in whole-mount Xenopus embryos. Genes expressed at neural plate stages (stages 14â18) are shown in panels AâL, and those only expressed at later stages are shown at mid/late tail bud stage (stages 28â32) in panels MâT. Several of the genes surveyed (Lhx5, Pou3f2b, Tbx15, Tbx6, Emx1.2 and Sim1 (AâD, M and N), were not expressed in the PPE, nor any placodal derivatives in later stages. Instead, such genes were expressed in the adjacent neural folds (Lhx5, Pou3f2b and Tbx15), ectoderm (Tbx6), or in the forebrain at later stages (Emx1.2). Several other genes were expressed broadly across the cranial ectoderm, at least partially overlapping with the PPE at neural plate stages (Ripply3, Crem, FosB and Znf214; E,OâQ), some of which are also maintained in placodal derivatives such as Znf214 in the otic vesicle (Q). The remaining genes (FâL, RâT) are expressed in parts of the PPE and maintained in some placodes (Hes2, Hes8, Hes9, Mab21l2b, Sox21, Isl2, Pou4f1.2, and Tlx1) or are expressed in a subset of placodes only (Atoh1, Gfi1a) (see Figure 4âfigure supplements 1â4 for additional stages). Yellow arrows mark placodal expression. Arrowheads mark non-placodal expression. Abbreviations: pA: anterior placodal region; pAD: anterior lateral line placode; pE: epibranchial placode; pL: lens placode; L: lens; pM: middle lateral line placode; pO: olfactory placode; pP: posterior placodal region; pPrV: profundal/trigenimal placodes; vOt: otic vesicle. Plots U and V show qPCR after Six1 or Eya1 overexpression. Log2 fold change values were calculated from qPCR data obtained after overexpression of Six1-GR (U) or Eya1-GR (V) in placodal explants and are shown next to corresponding fold change values obtained from the RNA-Seq data. In all cases shown, qPCR values broadly corroborate those from the RNA-Seq data - showing up-regulation of target genes after either Six1 or Eya1 overexpression. Vertical error bars show the standard deviation of the mean of biological triplicates.
Image published in: Riddiford N and Schlosser G (2016)
© 2016, Riddiford et al. Creative Commons Attribution license
| Gene | Synonyms | Species | Stage(s) | Tissue |
|---|---|---|---|---|
| lhx5.L | xlim-2a, xlim-5, Xlim5 | X. laevis | Throughout NF stage 14 | pre-chordal neural plate anterior placodal area |
| ripply3.L | LOC100145593, Xdscr6, XRipply3 | X. laevis | Throughout NF stage 14 | posterior placodal area chordal neural plate border |
| hes2.L | XHes2 | X. laevis | Throughout NF stage 14 | posterior placodal area neural fold neural plate border chordal neural plate border |
| isl2.L | islet-2, islet2b, LOC108711275 | X. laevis | Throughout NF stage 14 | anterior neural ridge anterior placodal area neurogenic placode |
| pou4f4.L | Brn3d, pou4f1.2 | X. laevis | Throughout NF stage 14 | posterior placodal area chordal neural plate border caudal presomitic mesoderm presomitic mesoderm |
| pou3f2.L | pou3f2-A, pou3f2-a, pou3f2-b, XlPOU3, XLPOU3b | X. laevis | Throughout NF stage 18 | neural plate pre-chordal neural plate chordal neural plate |
| tbx15.L | X. laevis | Throughout NF stage 18 | neural plate border pre-chordal neural plate neural fold anterior neural fold | |
| tbx6.L | XTbx6 | X. laevis | Throughout NF stage 18 | chordal neural plate chordal neural plate border caudal presomitic mesoderm presomitic mesoderm |
| hes5.4.L | hes8 | X. laevis | Throughout NF stage 18 | anterior placodal area neurogenic placode chordal neural plate chordal neural plate border anterior neural ridge neural fold anterior neural fold |
| hes5.7.L | esr9, hes9, hes9.1-a, hes9.1-b | X. laevis | Throughout NF stage 18 | anterior placodal area anterior neural ridge neurogenic placode chordal neural plate chordal neural plate border anterior neural fold neural fold |
| sox21.L | X. laevis | Throughout NF stage 18 | neural plate pre-chordal neural plate chordal neural plate | |
| mab21l2.L | mab21l2-a, mab21l2-b, Mab21l2b, Xmab-21, xmab21, xmab21l2 | X. laevis | Sometime during NF stage 18 to NF stage 19 | lens placode preplacodal ectoderm |
| emx1.L | empty spiracles homolog 1 | X. laevis | Throughout NF stage 28 | forebrain telencephalon |
| sim1.L | Xsim, Xsim1 | X. laevis | Throughout NF stage 28 | anterior |
| atoh1.L | ath1, bhlha14, hath1, math-1, math1, xath-1 | X. laevis | Sometime during NF stage 28 to NF stage 32 | otic vesicle hindbrain anterior lateral line placode |
| crem.L | X. laevis | Throughout NF stage 32 | neural crest cranial neural crest | |
| fosb.L | X. laevis | Throughout NF stage 32 | head region head neural crest cranial neural crest | |
| gfi1.L | gfi1a | X. laevis | Throughout NF stage 32 | lateral line ganglion otic vesicle ventral blood island posterior ventral blood island |
| tlx1.L | hox11, hox11-a, LOC108697427, Xhox11, XHox11L2 | X. laevis | Throughout NF stage 32 | lateral line ganglion otic vesicle profundus ganglion trigeminal ganglion posterior lateral line ganglion cement gland primary heart field secondary heart field |
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