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Multipotent progenitor populations are necessary for generating diverse tissue types during embryogenesis. We show the RNA polymerase-associated factor 1 complex (Paf1C) is required to maintain multipotent progenitors of the neural crest (NC) lineage in zebrafish. Mutations affecting each Paf1C component result in near-identical NC phenotypes; alyron mutant embryos carrying a null mutation in paf1 were analyzed in detail. In the absence of zygotic paf1 function, definitive premigratory NC progenitors arise but fail to maintain expression of the sox10 specification gene. The mutant NC progenitors migrate aberrantly and fail to differentiate appropriately. Blood and germ cell progenitor development is affected similarly. Development of mutant NC could be rescued by additional loss of positive transcription elongation factor b (P-TEFb) activity, a key factor in promoting transcription elongation. Consistent with the interpretation that inhibiting/delaying expression of some genes is essential for maintaining progenitors, mutant embryos lacking the CDK9 kinase component of P-TEFb exhibit a surfeit of NC progenitors and their derivatives. We propose Paf1C and P-TEFb act antagonistically to regulate the timing of the expression of genes needed for NC development., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)

The adenomatous polyposis coli (APC) tumor-suppressor protein, together with Axin and GSK3beta, forms a Wnt-regulated signaling complex that mediates phosphorylation-dependent degradation of beta-catenin by the proteasome. Siah-1, the human homolog of Drosophila seven in absentia, is a p53-inducible mediator of cell cycle arrest, tumor suppression, and apoptosis. We have now found that Siah-1 interacts with the carboxyl terminus of APC and promotes degradation of beta-catenin in mammalian cells. The ability of Siah-1 to downregulate beta-catenin signaling was also demonstrated by hypodorsalization of Xenopus embryos. Unexpectedly, degradation of beta-catenin by Siah-1 was independent of GSK3beta-mediated phosphorylation and did not require the F box protein beta-TrCP. These results indicate that APC and Siah-1 mediate a novel beta-catenin degradation pathway linking p53 activation to cell cycle control.

The vertebrate body plan has bilateral symmetry and left-right asymmetries that are highly conserved. The molecular pathways for left-right development are beginning to be elucidated. Several distinct mechanisms to initiate the vertebrate left-right axis have been proposed. These mechanisms appear to converge on highly conserved expression patterns of genes in the transforming growth factor-beta (TGFbeta) family of cell-cell signaling factors, nodal and lefty-2, and subsequently the expression of the transcription regulator Pitx2, in left lateral plate mesoderm. It is possible that downstream signaling pathways diverge in distinct classes of vertebrates.

The embryonic midline in vertebrates has been implicated in left-right development, but the mechanisms by which it regulates left-right asymmetric gene expression and organ morphogenesis are unknown. Zebrafish embryos have three domains of left-right asymmetric gene expression that are useful predictors of organ situs. cyclops (nodal), lefty1 and pitx2 are expressed in the left diencephalon; cyclops, lefty2 and pitx2 are expressed in the left heart field; and cyclops and pitx2 are expressed in the left gut primordium. Distinct alterations of these expression patterns in zebrafish midline mutants identify four phenotypic classes that have different degrees of discordance among the brain, heart and gut. These classes help identify two midline domains and several genetic pathways that regulate left-right development. A cyclops-dependent midline domain, associated with the prechordal plate, regulates brain asymmetry but is dispensable for normal heart and gut left-right development. A second midline domain, associated with the anterior notochord, is dependent on no tail, floating head and momo function and is essential for restricting asymmetric gene expression to the left side. Mutants in spadetail or chordino give discordant gene expression among the brain, heart and gut. one-eyed pinhead and schmalspur are necessary for asymmetric gene expression and may mediate signaling from midline domains to lateral tissues. The different phenotypic classes help clarify the apparent disparity of mechanisms proposed to explain left-right development in different vertebrates.

The Lefty subfamily of TGFbeta signaling molecules has been implicated in early development in mouse, zebrafish, and chick. Here, we show that Xenopus lefty (Xlefty) is expressed both bilaterally in symmetric midline domains and unilaterally in left lateral plate mesoderm and anterior dorsal endoderm. To examine the roles of Xlefty in left-right development, we created a system for scoring gut asymmetry and examined the effects of unilateral Xlefty misexpression on gut development, heart development, and Xnr-1 and XPitx2 expression. In contrast to the unilateral effects of Vg1, Activin, Nodal, or BMPs, targeted expression of Xlefty in either the left or the right side of Xenopus embryos randomized the direction of heart looping, gut coiling, and left-right positioning of the gut and downregulated the asymmetric expression of Xnr-1 and XPitx2. It is currently thought that Lefty proteins act as feedback inhibitors of Nodal signaling. However, this would not explain the effects of right-sided Xlefty misexpression. Here, we show that Xlefty interacts with the signaling pathways of other members of the TGFbeta family during left-right development. Results from coexpression of Xlefty and Vg1 indicate that Xlefty can nullify the effects of Vg1 ectopic expression and that Xlefty is downstream of left-sided Vg1 signaling. Results from coexpression of Xlefty and XBMP4 indicate that XLefty and XBMP4 interact both synergistically and antagonistically in a context-dependent manner. We propose a model in which interactions of Xlefty with multiple members of the TGFbeta family enhance the differences between the right-sided BMP/ALK2/Smad pathway and the left-sided Vg1/anti-BMP/Nodal pathway, leading to left-right morphogenesis of the gut and heart., (Copyright 2000 Academic Press.)

The pitx2 gene is a member of the bicoid-homeodomain class of transcription factors that has been implicated in the control of left-right asymmetry during organogenesis. Here we demonstrate that in zebrafish there are two pitx2 isoforms, pitx2a and pitx2c, which show distinct expression patterns and have non-overlapping functions during mesendoderm and asymmetric organ development. pitx2c is expressed symmetrically in presumptive mesendoderm during late blastula stages and in the prechordal plate during late gastrulation. pitx2a expression is first detected at bud stage in the anterior prechordal plate. The regulation of early mesendoderm pitx2c expression is dependent on one-eyed pinhead (EGF-CFC-related gene) and spadetail (tbx-transcription factor) and can be induced by ectopic goosecoid expression. Maintenance of pitx2c midline expression is dependent on cyclops (nodal) and schmalspur, but not no tail (brachyury). Ectopic expression of pitx2 isoforms results in distinct morphological and molecular phenotypes, indicating that pitx2a and pitx2c have divergent regulatory functions. Both isoforms downregulate goosecoid on the dorsal side, but in contrast to earlier reports that nodal and lefty are upstream of pitx2, ectopic pitx2c in other regions induces cyclops, lefty2 and goosecoid expression. Asymmetric isoform expression occurs in non-overlapping domains, with pitx2c in left dorsal diencephalon and developing gut and pitx2a in left heart primordium. Targeted asymmetric expression in Xenopus shows that both isoforms can alter left-right development, but pitx2a has a slightly stronger effect on heart laterality. Our results indicate that distinct genetic pathways regulate pitx2a and pitx2c isoform expression, and each isoform regulates different downstream pathways during mesendoderm and asymmetric organ development.