Expression of achaete-scute homolog 3 in Xenopus embryos converts ectodermal cells to a neural fate.

Abstract

In Drosophila, the proneural genes of the achaete-scute complex encode transcriptional activators that can commit cells to a neural fate. We have isolated cDNAs for two Xenopus achaete-scute homologs, ASH3a and ASH3b, which are expressed in a subset of central nervous system (CNS) neuroblasts during early neurogenesis. After expressing either ASH3 protein in developing Xenopus embryos, we find enlargement of the CNS at the expense of adjacent non-neural ectoderm. Analysis of molecular markers for neural, epidermal, and neural crest cells indicates that CNS expansion occurs as early as neural plate formation. ASH3-dependent CNS enlargement appears to require neural induction, as it does not occur in animal cap explants. Inhibition of DNA synthesis shows that additional CNS tissue does not depend on cell division--rather it reflects conversion of prospective neural crest and epidermal cells to a neural fate. The differentiation of the early forming primary neurons also seems to be prevented by ASH3 expression. This may be secondary to the observed activation of Xotch transcription by ASH3.

Novel functions for Period 3 and Exo-rhodopsin in rhythmic transcription and melatonin biosynthesis within the zebrafish pineal organ

“…The mMessage Machine Kit (Ambion Inc. Austin, Tx, USA) was used to synthesize capped mRNA in vitro from the pCS2+otx5, pCS2+β-gal, and pCS2+ RFP plasmids [20,48,57]. Single blastomeres of 8-16 cell stage exorh:GFP embryos were injected with 0.5-1.5 nl of mRNA solution using a PLI-90 injector.…”

Improved translation efficiency of injected mRNA during early embryonic development

“…The pCS2 vector (Rupp et al, 1994;Turner and Weintraub, 1994) represents another backbone frequently used for misexpression experiments. Of interest, we observed an almost 20-fold increase in luciferase activity for mRNA derived from this construct compared with the KW luc reference ( Table 2).…”

Xath5 regulates neurogenesis in the Xenopus olfactory placode

“…Xath5 overexpression causes expansion of Xebf2 expression in the presence of HUA, which blocks cell division, arguing that surrounding ectodermal cells are being recruited to the ORN fate in response to Xath5 expression. The proneural bHLH factors have been shown to convert ectodermal cells, which would normally differentiate as epidermis, into neurons, when overexpressed in the developing Xenopus embryo (Ferreiro et al, 1994;Turner and Weintraub, 1994;Ma et al, 1996;Lee, 1997b). Indeed, overexpression of Xath5 promotes the formation of ectopic N-tubulin-positive neurons throughout the ectoderm of injected embryos (Kanekar et al, 1997).…”

“…Embryos were allowed to develop then collected at various stages according to Nieuwkoop and Faber (1994). Embryos were fixed in MEMFA (Harland et al, 1991) at room temperature for less than 1 hr and stained with X-gal by using previously described methods (Turner and Weintraub, 1994), stored in methanol and analyzed by means of in situ hybridization.…”

“…Embryos were collected at stage 24 -26 and fixed for 30 -60 min in MEMFA. Embryos were subsequently stained with X-gal as previously described (Turner and Weintraub, 1994) and then processed by in situ hybridization for Xebf2 expression.…”