Interactions between Wnt and Vg1 signalling pathways initiate primitive streak formation in the chick embryo.

The posterior marginal zone (PMZ) of the chick embryo has Nieuwkoop centre-like properties: when transplanted to another part of the marginal zone, it induces a complete embryonic axis, without making a cellular contribution to the induced structures. However, when the PMZ is removed, the embryo can initiate axis formation from another part of the remaining marginal zone. Chick Vg1 can mimic the axis-inducing ability of the PMZ, but only when misexpressed somewhere within the marginal zone. We have investigated the properties that define the marginal zone as a distinct region. We show that the competence of the marginal zone to initiate ectopic primitive streak formation in response to cVg1 is dependent on Wnt activity. First, within the Wnt family, only Wnt8C is expressed in the marginal zone, in a gradient decreasing from posterior to anterior. Second, misexpression of Wnt1 in the area pellucida enables this region to form a primitive streak in response to cVg1. Third, the Wnt antagonists Crescent and Dkk-1 block the primitive streak-inducing ability of cVg1 in the marginal zone. These findings suggest that Wnt activity defines the marginal zone and allows cVg1 to induce an axis. We also present data suggesting some additional complexity: first, the Vg1 and Wnt pathways appear to regulate the expression of downstream components of each other's pathway; and second, misexpression of different Wnt antagonists suggests that different classes of Wnts may cooperate with each other to regulate axis formation in the normal embryo.

Reconciling different models of forebrain induction and patterning: a dual role for the hypoblast.

Several models have been proposed for the generation of the rostral nervous system. Among them, Nieuwkoop's activation/transformation hypothesis and Spemann's idea of separate head and trunk/tail organizers have been particularly favoured recently. In the mouse, the finding that the visceral endoderm (VE) is required for forebrain development has been interpreted as support for the latter model. Here we argue that the chick hypoblast is equivalent to the mouse VE, based on fate, expression of molecular markers and characteristic anterior movements around the time of gastrulation. We show that the hypoblast does not fit the criteria for a head organizer because it does not induce neural tissue from naïve epiblast, nor can it change the regional identity of neural tissue. However, the hypoblast does induce transient expression of the early markers Sox3 and Otx2. The spreading of the hypoblast also directs cell movements in the adjacent epiblast, such that the prospective forebrain is kept at a distance from the organizer at the tip of the primitive streak. We propose that this movement is important to protect the forebrain from the caudalizing influence of the organizer. This dual role of the hypoblast is more consistent with the Nieuwkoop model than with the notion of separate organizers, and accommodates the available data from mouse and other vertebrates.

Induction of primitive streak and Hensen's node by the posterior marginal zone in the early chick embryo.

In the preprimitive streak chick embryo, the search for a region capable of inducing the organizer, equivalent to the Nieuwkoop Center of the amphibian embryo, has focused on Koller's sickle, the hypoblast and the posterior marginal zone. However, no clear evidence for induction of an organizer without contribution from the inducing tissue has been provided for any of these structures. We have used DiI/DiO labeling to establish the fate of midline cells in and around Koller's sickle in the normal embryo. In the epiblast, the boundary between cells that contribute to the streak and those that do not lies at the posterior edge of Koller's sickle, except at stage X when it lies slightly more posteriorly in the epiblast. Hypoblast and endoblast (a second lower layer formed under the streak) have distinct origins in the lower layer, and goosecoid expression distinguishes between them. We then used anterior halves of chick prestreak embryos as recipients for grafts of quail posterior marginal zone; quail cells can be identified subsequently with a quail-specific antibody. Anterior halves alone usually formed a streak, most often from the posterior edge. Quail posterior marginal zones without Koller's sickle were grafted to the anterior side of anterior halves. These grafts were able to increase significantly the frequency of streaks arising from the anterior pole of stage X-XI anterior halves without contributing to the streak or node. Stage XII anterior halves no longer responded. A goosecoid-expressing hypoblast did not form under the induced streak, indicating that it is not required for streak formation. We conclude that the marginal zone posterior to Koller's sickle can induce a streak and node, without contributing cells to the induced streak.

Misexpression of chick Vg1 in the marginal zone induces primitive streak formation.

In the chick embryo, the primitive streak is the first axial structure to develop. The initiation of primitive streak formation in the posterior area pellucida is influenced by the adjacent posterior marginal zone (PMZ). We show here that chick Vg1 (cVg1), a member of the TGFbeta family of signalling molecules whose homolog in Xenopus is implicated in mesoderm induction, is expressed in the PMZ of prestreak embryos. Ectopic expression of cVg1 protein in the marginal zone chick blastoderms directs the formation of a secondary primitive streak, which subsequently develops into an ectopic embryo. We have used cell marking techniques to show that cells that contribute to the ectopic primitive streak change fate, acquiring two distinct properties of primitive streak cells, defined by gene expression and cell movements. Furthermore, naive epiblast explants exposed to cVg1 protein in vitro acquire axial mesodermal properties. Together, these results show that cVg1 can mediate ectopic axis formation in the chick by inducing new cell fates and they permit the analysis of distinct events that occur during primitive streak formation.

Starvation stress modulates the expression of the Aspergillus nidulans brlA regulatory gene.

Expression of the Aspergillus nidulans brlA gene plays a fundamental role in the switch from vegetative growth to asexual reproduction. Using a media-shifting protocol to induce submerged sporulation and brlA-lacZ as an expression marker, it was shown that carbon and nitrogen starvation stress induced brlA transcription to different degrees. Glucose starvation induced briA rapidly to high levels and resulted in spore formation on reduced conidiophores, whereas nitrogen starvation induced brlA gradually to lower levels and sporulation occurred to a lesser extent but from more complex conidiophores. beta-Galactosidase activity paralleled brlA alpha and brlA beta mRNA. No clear qualitative differences between the two brlA transcripts were found in these starvation conditions, suggesting that the different patterns of sporulation could be explained by quantitative expression differences. Since brlA mRNA did not accumulate in the presence of a high glucose concentration, we investigated the role of other carbon sources on brlA expression. Non-repressing carbon sources such as glycerol, acetate and arabinose were as effective as glucose in preventing brlA mRNA accumulation, suggesting that the glucose effects on brlA expression could be explained as a response to nutrient starvation, rather than by carbon catabolite repression. Despite similar low levels of brlA transcripts being detected during growth in glucose or non-repressing carbon sources, conidiophores were formed only in medium containing glycerol, acetate or arabinose. When mycelia were not shifted to starvation conditions, sporulation was not observed in standard minimal medium even after glucose was exhausted, unless the medium was buffered.(ABSTRACT TRUNCATED AT 250 WORDS)