A time delay gene circuit is required for palp formation in the ascidian embryo

Abstract

The ascidian larval brain and palps (a putative rudimentary placode) are specified by two transcription factor genes, ZicL and FoxC, respectively. FGF9/16/20 induces ZicL expression soon after the bipotential ancestral cells divide into the brain and palp precursors at the early gastrula stage. FGF9/16/20 begins to be expressed at the 16-cell stage, and induces several target genes, including Otx, before the gastrula stage. Here, we show that ZicL expression in the brain lineage is transcriptionally repressed by Hes-a and two Blimp-1-like zinc finger proteins, BZ1 and BZ2, in the bi-potential ancestral cells. ZicL is precociously expressed in the bi-potential cells in embryos in which these repressors are knocked down. This precocious ZicL expression produces extra brain cells at the expense of palp cells. The expression of BZ1 and BZ2 is turned off by a negative autofeedback loop. This auto-repression acts as a delay circuit that prevents ZicL from being expressed precociously before the brain and palp fates split, thereby making room within the neural plate for the palps to be specified. Addition of the BZ1/2 delay timer circuit to the gene regulatory network responsible for brain formation might represent a key event in the acquisition of the primitive palps/placodes in an ancestral animal.

Differential temporal control of Foxa.a and Zic-r.b specifies brain versus notochord fate in the ascidian embryo

“…The notochord developmental program was ectopically activated in Prdm1-r.a/b/Hes.a morphants We previously showed that Zic-r.b is precociously expressed in bipotential brain/palp progenitors at the 64-cell stage in doublemorphant embryos of Prdm1-r.a and Prdm1-r.b (Prdm1-r.a/b morphants), and this ectopic activation of Zic-r.b converts palp fate into brain fate (Ikeda et al, 2013). We also showed that Zic-r.b expression begins earlier (at the 32-cell stage) in Prdm1-r.a/b/Hes.a triple morphants.…”

“…Three transcriptional repressors, Prdm1-r.a (formerly BZ1), Prdm1-r.b (formerly BZ2) and Hes.a are important for ensuring that Foxa.a and Zic-r.b are expressed sequentially in the presumptive brain cells, as indicated by the fact that Zic-r.b is precociously expressed at the 32-cell stage in triple morphants of Prdm1-r.a, Prdm1-r.b and Hes.a (Prdm1-r.a/b/Hes.a morphants) (Ikeda et al, 2013). In the present study, we show that Prdm1-r.a also promotes termination of Foxa.a expression, and we propose a robust mechanism for temporally distinct expression of Foxa.a and Zic-r.b to ensure the brain lineage specification.…”

Islet is a key determinant of ascidian palp morphogenesis

“…Btd (also known as Sp8 or ZF220) is expressed in the palp lineage at the early tailbud stage, downstream of FoxC, the earliest marker of the palp lineage (Ikeda et al, 2013;Imai et al, 2004Imai et al, , 2006. In late tailbud embryos, Btd appears to be co-expressed with Emx in the rings, and also occurs in the intervening anterior-most ectoderm, but is specifically excluded from the center of the rings (Fig.…”

“…Specifically, we showed that perturbation of FGF-MAPK signaling led to ectopic FoxC expression, but larvae that developed under these conditions failed to develop palps. A recent study revealed that, whereas FoxC is a marker of the palp lineage, it is not a marker of palp fate; FoxC expression in the palp lineage persists under conditions that inhibit palp development (Ikeda et al, 2013). Moreover, Islet expression is lost upon MAPK inhibition from the 8-cell stage onward, and MAPK signaling through the neurula stage is reported to be required for normal palp development (Hudson et al, 2003).…”

“…S1 for the complete sequence. The Islet DNA-binding domain was amplified with Isl DBD NheF: TAATGCTAG-CTAAAGATGCGAAGACGACGCGAG and DBD NgoMIV R: TAATGC-CGGCCTTCGCTTGCTGCTCCTGGATTTG, and cloned upstream of the Based on results of current and previous studies (Ikeda et al, 2013;Imai et al, 2006;Wagner and Levine, 2012). Genes in bold black font are expressed in indicated region; gray font indicates genes repressed in those areas; blue font denotes untested regulatory interactions; parentheses indicate genes that were expressed in progenitors of that region but have since been downregulated.…”