Uwe Walldorf

An urbilaterian origin of the tripartite brain: developmental genetic insights from Drosophila.

Studies on expression and function of key developmental control genes suggest that the embryonic vertebrate brain has a tripartite ground plan that consists of a forebrain/midbrain, a hindbrain and an intervening midbrain/hindbrain boundary region, which are characterized by the specific expression of the Otx, Hox and Pax2/5/8 genes, respectively. We show that the embryonic brain of the fruitfly Drosophila melanogaster expresses all three sets of homologous genes in a similar tripartite pattern. Thus, a Pax2/5/8 expression domain is located at the interface of brain-specific otd/Otx2 and unpg/Gbx2 expression domains anterior to Hox expression regions. We identify this territory as the deutocerebral/tritocerebral boundary region in the embryonic Drosophila brain. Mutational inactivation of otd/Otx2 and unpg/Gbx2 result in the loss or misplacement of the brain-specific expression domains of Pax2/5/8 and Hox genes. In addition, otd/Otx2 and unpg/Gbx2 appear to negatively regulate each other at the interface of their brain-specific expression domains. Our studies demonstrate that the deutocerebral/tritocerebral boundary region in the embryonic Drosophila brain displays developmental genetic features similar to those observed for the midbrain/hindbrain boundary region in vertebrate brain development. This suggests that a tripartite organization of the embryonic brain was already established in the last common urbilaterian ancestor of protostomes and deuterostomes.

Differential expression and function of the Drosophila Pax6 genes eyeless and twin of eyeless in embryonic central nervous system development

We analyzed the expression and function of eyeless (ey) and twin of eyeless (toy) in the embryonic central nervous system (CNS) of Drosophila. Both genes are differentially expressed in specific neuronal subsets (but not in glia) in every CNS neuromere, and in the brain, specific cell populations co-expressing both proteins define a longitudinal domain which is intercalated between broad exclusive expression domains of ey and toy. Studies of genetic null alleles and dsRNA interference did not reveal any gross neuroanatomical effects of ey, toy, or ey/toy elimination in the embryonic CNS. In contrast, targeted misexpression of ey, but not of toy, resulted in profound axonal abnormalities in the embryonic ventral nerve cord and brain.

Expression, regulation and function of the homeobox gene empty spiracles in brain and ventral nerve cord development of Drosophila.

We analyse the role of the empty spiracles (ems) gene in embryonic brain and ventral nerve cord development. ems is differentially expressed in the neurectoderm of the anterior head versus the trunk region of early embryos. A distal enhancer region drives expression in the deutocerebral brain anlage and a proximal enhancer region drives expression in the VNC and tritocerebral brain anlage. Mutant analysis indicates that in the anterior brain ems is necessary for regionalized neurogenesis in the deutocerebral and tritocerebral anlagen. In the posterior brain and VNC ems is necessary for correct axonal pathfinding of specific interneurons. Rescue experiments indicate that the murine Emx2 gene can partially replace the fly ems gene in CNS development.

Interaction of gap genes in the Drosophila head: tailless regulates expression of empty spiracles in early embryonic patterning and brain development

Unlike gap genes in the trunk region of Drosophila embryos, gap genes in the head were presumed not to regulate each others transcription. Here, we show that in tailless (tll) loss-of-function mutants the empty spiracles (ems) expression domain in the head expands, whereas it retracts in tll gain-of-function embryos. We have identified a 304bp element in the ems-enhancer which is sufficient to drive expression in the head and brain and which contains two TLL and two BCD binding sites. Transgenic reporter gene lines containing mutations of the TLL binding sites demonstrate that tll directly inhibits the expression of ems in the early embryonic head and the protocerebral brain anlage. These results are the first demonstration of direct transcriptional regulation between gap genes in the head.

Patterning defects in the primary axonal scaffolds caused by the mutations of the extradenticle and homothorax genes in the embryonic Drosophila brain

Abstract During early brain development in Drosophila a highly stereotyped pattern of axonal scaffolds evolves by precise pioneering and selective fasciculation of neural fibers in the newly formed brain neuromeres. Using an axonal marker, Fasciclin II, we show that the activities of the extradenticle(exd) and homothorax (hth) genes are essential to this axonal patterning in the embryonic brain. Both genes are expressed in the developing brain neurons, including many of the tract founder cluster cells. Consistent with their expression profiles, mutations of exd and hth strongly perturb the primary axonal scaffolds. Furthermore, we show that mutations of exd and hth result in profound patterning defects of the developing brain at the molecular level including stimulation of the orthodenticle gene and suppression of the empty spiracles and cervical homeotic genes. In addition, expression of a Drosophila Pax6 gene, eyeless, is significantly suppressed in the mutants except for the most anterior region. These results reveal that, in addition to their homeotic regulatory functions in trunk development, exd and hth have important roles in patterning the developing brain through coordinately regulating various nuclear regulatory genes, and imply molecular commonalities between the developmental mechanisms of the brain and trunk segments, which were conventionally considered to be largely independent.

Homeobrain, a novel paired-like homeobox gene is expressed in the Drosophila brain

The homeobrain (hbn) gene is a new paired-like homeobox gene which is expressed in the embryonic brain and the ventral nerve cord. Expression of homeobrain initiates during the blastoderm stage in the anterior dorsal head primordia and the gene is persistently expressed in these cells which form parts of the brain during later embryonic stages. An additional weaker expression pattern is detected in cells of the ventral nerve cord from stage 11 on. The homeodomain in the Homeobrain protein is most similar to the Drosophila proteins DRx, Aristaless and Munster. In addition, the localized brain expression patterns of homeobrain and DRx resemble each other. Two other homeobox genes, orthopedia and DRx are clustered in the 57B region along with homeobrain. The current evidence indicates that homeobrain, DRx and orthopedia form a homeobox gene cluster in which all the members are expressed in specific embryonic brain subregions.

Development of the Deformed protein pattern in the embryo of the honeybee Apis mellifera L. (Hymenoptera)

SummaryWe have raised antiserum against part of the Deformed (Dfd) protein of the honeybee and describe here the expression pattern of the Dfd protein during honeybee embryogenesis. Dfd protein is first stained in the prospective gnathal region of the cellular blastoderm. This circumferential band corresponds to the distribution of Dfd mRNA described earlier, and to the blastodermal Dfd expression pattern in Drosophila. Using an antibody against the engrailed (en) protein of Drosophila, we found that at the beginning of gastrulation Dfd expression in the honeybee, as in Drosophila, is restricted to the future intercalary, mandibular and maxillary segments. During gastrulation, the mesodermal nuclei loose the Dfd label gradually from anterior to posterior, and in the ectoderm the most posterior ventral cells loose Dfd while retaining en staining; thus, in contrast to what has been described for Drosophila, the posterior Dfd expression border seems to move forward ventrally to the parasegmental boundary within the maxillary segment. In the late germ band, the lateral tips of the Dfd-expressing band are connected across the dorsal side by a row of amnion cells with strongly staining large nuclei. After dorsal closure, a narrow stripe of Dfd-staining dorsal cells behind the neck region may indicate that the maxillary segment contributes to the dorsal body wall posterior to the head capsule. Thus, apart from some minor deviations, the Dfd expression pattern in the honeybee strongly resembles that in Drosophila prior to head involution. This is compatible with the assumption that head involution (which is a special adaption in higher dipterans) ensues after a rather conserved course of early head development in which Dfd appears to play a basic role.

In situ localization of the transcripts of a homeobox gene in the honeybee Apis mellifera L. (Hymenoptera)

SummaryWe have isolated and characterized a homeoboxcontaining gene from the honeybee Apis mellifera. Its homeobox region shows a high degree of sequence similarity to the homeobox of the Drosophila gene Deformed (Dfd). At the DNA level 82% of the basepairs are the same, whereas the putative amino acid sequences are identical between the bee and the fruitfly genes. Similarity is also present 5′ and 3′ to the homeobox. Using this isolate as a probe we have performed in situ hybridization on sections from blastoderm-stage embryos of the honeybee Apis mellifera. In early blastoderm stages we found a rather irregular pattern of labelled nuclei. In middle stages we found silver grains over each nucleus and also over the cytoplasm in a belt of blastoderm cells in the prospective gnathal region. These results indicate that the Deformed genes from honeybee and fruitfly are homologous both with respect to their DNA sequence and their spatial and temporal pattern of expression during embryogenesis.