Kegg Pathway: Dorso-ventral axis formation

Dev Dyn. 2009 Aug 13;
Ulloa F, Martí E

The spinal cord has been used as a model to dissect the mechanisms that govern the patterning of tissues during animal development, since the principles that rule the Dorso-ventral patterning of the neural tube are applicable to other systems. Signals that determine the Dorso-ventral axis of the spinal cord include Sonic hedgehog (Shh), acting as a bona fide morphogenetic signal to determine ventral progenitor identities, and members of the Bmp and the Wnt families, acting in the dorsal neural tube. Although Wnts have been initially recognized as important in proliferation of neural progenitor cells, their role in the Dorso-ventral patterning has been controversial. In this review, we discuss recent reports that show an important contribution of the Wnt canonical pathway in Dorso-ventral pattern formation. These data allow building a model by which the ventralizing activity of Shh is antagonized by Wnt activity through the expression of Gli3, a potent inhibitor of the Shh pathway. Therefore, antagonistic interactions between canonical Wnt, promoting dorsal identities, and Shh pathways, inducing ventral ones, would define the Dorso-ventral patterning of the developing central nervous system. Developmental Dynamics, 2009. (c) 2009 Wiley-Liss, Inc.

Nat Cell Biol. 2009 Sep; 11(9): 1121-7
Ro H, Dawid IB

The organizer anchors the primary embryonic axis, and balance between dorsal (organizer) and ventral domains is fundamental to body patterning. LNX (ligand of Numb protein-X) is a RING finger and four PDZ domain-containing E3 ubiquitin ligase. LNX serves as a binding platform and may have a role in cell fate determination, but its in vivo functions are unknown. Here we show that Lnx-l (Lnx-like) functions as a critical regulator of Dorso-ventral axis formation in zebrafish. Depletion of Lnx-l using specific antisense morpholinos (MOs) caused strong embryonic dorsalization. We identified Bozozok (Boz, also known as Dharma or Nieuwkoid) as a binding partner and substrate of Lnx-l. Boz is a homeodomain-containing transcriptional repressor induced by canonical Wnt signalling that is critical for dorsal organizer formation. Lnx-l induced K48-linked polyubiquitylation of Boz, leading to its proteasomal degradation in human 293T cells and in zebrafish embryos. Dorsalization induced by Boz overexpression was suppressed by raising the level of Lnx-l, but Lnx-l failed to counteract dorsalization caused by mutant Boz lacking a critical motif for Lnx-l binding. Furthermore, dorsalization induced by depletion of Lnx-l was alleviated by attenuation of Boz expression. We conclude that Lnx-l modulates Boz activity to prevent the invasion of ventral regions of the embryo by organizer tissue. These studies introduce a ubiquitin ligase, Lnx-l, as a balancing modulator of axial patterning in the zebrafish embryo.

PLoS Biol. 2009 Jun 16; 7(6): e1000135
Zlatic M, Li F, Strigini M, Grueber W, Bate M

During the development of neural circuitry, neurons of different kinds establish specific synaptic connections by selecting appropriate targets from large numbers of alternatives. The range of alternative targets is reduced by well organised patterns of growth, termination, and branching that deliver the terminals of appropriate pre- and postsynaptic partners to restricted volumes of the developing nervous system. We use the axons of embryonic Drosophila sensory neurons as a model system in which to study the way in which growing neurons are guided to terminate in specific volumes of the developing nervous system. The mediolateral positions of sensory arbors are controlled by the response of Robo receptors to a Slit gradient. Here we make a genetic analysis of factors regulating position in the Dorso-ventral axis. We find that Dorso-ventral layers of neuropile contain different levels and combinations of Semaphorins. We demonstrate the existence of a central to dorsal and central to ventral gradient of Sema 2a, perpendicular to the Slit gradient. We show that a combination of Plexin A (Plex A) and Plexin B (Plex B) receptors specifies the ventral projection of sensory neurons by responding to high concentrations of Semaphorin 1a (Sema 1a) and Semaphorin 2a (Sema 2a). Together our findings support the idea that axons are delivered to particular regions of the neuropile by their responses to systems of positional cues in each dimension.

Curr Opin Genet Dev. 2009 Aug; 19(4): 404-11
Fonseca RN, Lynch JA, Roth S

Drosophila melanogaster has been used as an arthropod model to understand the establishment of embryonic axes. However, the long germ type of embryogenesis in Drosophila is highly divergent from the ancestral, short germ mode seen in most other arthropods. Therefore, broader sampling of other arthropod taxa is required to understand the evolution of axial patterning mechanisms within this phylum. Here we describe recent progress toward this end. First, we describe differing strategies for providing patterning information along the anterior-posterior axis of arthropod embryos. Second, we discuss the recent findings on Dorso-ventral axis of beetles and spiders that highlight the importance of regulatory interactions at the zygotic level. Third, recent discoveries of the roles of Wnt signalling, the Notch/Delta pathway and caudal in establishing and patterning the posterior embryonic region in spiders and beetles are described. In the end, we try to integrate these new findings with non-model arthropods in a common framework and discuss the major gaps of knowledge that should be addressed by future studies.

PLoS Biol. 2009 May; 7(5): e1000101
Harvey SA, Smith JC

During embryonic development, signalling molecules known as morphogens act in a concentration-dependent manner to provide positional information to responding tissues. In the early zebrafish embryo, graded signalling by members of the nodal family induces the formation of mesoderm and endoderm, thereby patterning the embryo into three germ layers. Nodal signalling has also been implicated in the establishment of the Dorso-ventral axis of the embryo. Although one can infer the existence of nodal gradients by comparing gene expression patterns in wild-type embryos and embryos in which nodal signalling is diminished or augmented, real understanding can only come from directly observing the gradients. One approach is to determine local ligand concentrations in the embryo, but this is technically challenging, and the presence of inhibitors might cause the effective concentration of a ligand to differ from its actual concentration. We have therefore taken two approaches to visualise a direct response to nodal signalling. In the first, we have used transgenic embryos to study the nuclear accumulation of a Smad2-Venus fusion protein, and in the second we have used bimolecular fluorescence complementation to visualise the formation of a complex between Smad2 and Smad4. This has allowed us to visualise, in living embryos, the formation of a graded distribution of nodal signalling activity. We have quantified the formation of the gradient in time and space, and our results not only confirm that nodal signalling patterns the embryo into three germ layers, but also shed light on its role in patterning the Dorso-ventral axis and highlight unexpected complexities of mesodermal patterning.

C R Biol. 2009 Feb-Mar; 332(2-3): 210-8
Coolen M, Menuet A, Mazan S

The genetic mechanisms, which control axis specification, apparently extensively diverge across vertebrates. In amphibians and teleosts, they are tightly linked to the establishment of an early Dorso-ventral polarity. This polarity has no equivalent in amniotes, which unlike the former, retain a considerable plasticity for their site of axis formation until blastula stages and rely on signals secreted by extra-embryonic tissues for the establishment of their early rostro-caudal pattern. In order to better understand the links between these seemingly highly divergent mechanisms, we have used an evo-devo approach, aimed at reconstructing the gnathostome ancestral state and focussed on a chondrichthyan, the dogfish Scyliorhinus canicula. A detailed molecular characterization of the dogfish embryo at blastula and gastrula stages highlights striking similarities with all vertebrate model organisms including amniotes. It suggests the presence in the dogfish of territories homologous to the hypoblast and extra-embryonic ectoderm of the latter, which may therefore reflect the primitive condition of jawed vertebrates. In the ancestral state, these territories are specified at opposite sides of an early axis of bilateral symmetry, homologous to the Dorso-ventral axis of amphibians and teleosts, and aligned with the later forming embryonic axis, from head to tail. Amniotes have diverged from this pattern through a posterior expansion of extra-embryonic ectoderm, resulting in an apparently radial symmetry at late blastula stages. These data delineate the broad outlines of the gnathostome ancestral pattern of axis specification and highlight an unexpected unity of mechanisms across jawed vertebrates. They illustrate the complementarity of comparative and genetic approaches for a comprehensive view of developmental mechanisms themselves.

Dev Dyn. 2009 Jan; 238(1): 194-203
Illes JC, Winterbottom E, Isaacs HV

Gsx class proteins are members of the ParaHox homeodomain transcription factor family with conserved roles in specification and patterning of the nervous system. We report the cloning of two Gsx genes, Gsh1 and Gsh2, from the frog Xenopus tropicalis. We demonstrate the existence of a single, intact Xenopus ParaHox cluster, containing Gsh1, Pdx, and Cdx2, plus three degenerate clusters containing Gsh2, Cdx1, and Cdx4. Anterior expression boundaries of genes from the intact ParaHox cluster are co-linear with respect to their genomic organization. We show that Gsh1 and Gsh2 exhibit complex, overlapping patterns of expression within the anterior nervous system from open neural plate stages. We also find that expression of Gsh2, Nkx6, and Msx1 across the medio-lateral axis of the amphibian neural plate is strikingly similar to that of related genes in the Drosophila neuroectoderm. These findings provide further evidence for a conserved pathway regulating Dorso-ventral patterning in the Bilateria.

Gene Expr Patterns. 2009 Feb; 9(2): 94-108
Bethea CL, Reddy AP, Pedersen D, Tokuyama Y

The rhesus monkey embryonic stem cell line 366.4 differentiates into serotonin neurons. We examined the genetic cascade during differentiation and compared ESC-derived serotonin neurons to adult monkey serotonin neurons. RNA was extracted from ESC colonies, embryoid bodies (EBs), neurospheres in selection (N1) and proliferation stages (N2), differentiated serotonin neurons (N3) and from laser captured (LC) serotonin neurons of spayed female macaques treated with placebo, estrogen (E), progesterone (P) or E+P. The RNA was labeled and hybridized to Rhesus Monkey Affymetrix Gene Chips (n=1 per stage and 2 per animal treatment). Gene expression was examined with GeneSifter software. 545 genes that were related to developmental processes showed a threefold or greater change between stages. TGFb, Wnt, VEGF and Hedgehog signaling pathways showed the highest percent of probe set changes during differentiation. Genes in the categories (a) homeobox binding and transcription factors, (b) growth factors and receptors, (c) brain and neural specific factors and (d) serotonin specific factors are reported. Pivotal genes were confirmed with quantitative RT-PCR. In the serotonin developmental cascade, FGFR2 was robustly expressed at each stage. GATA3 was robustly expressed in EBs. Sonic hedgehog (Shh), PTCH (Shh-R) and Fev1 transcription factor expression coincided with the induction of serotonin specific marker genes during N1-selection. A majority of the examined genes were expressed in adult serotonin neurons. However, in the ESC-derived neurons, there was significant over-representation of probe sets related to cell cycle, axon guidance & Dorso-ventral axis formation. This analysis suggests that the 366.4 cell line possesses cues for serotonin differentiation at early stages of differentiation, but that ESC-derived serotonin neurons are still immature.

Int J Dev Biol. 2008; 52(7): 857-71
Fernandez-Teran M, Ros MA

The Apical Ectodermal Ridge (AER) is one of the main signaling centers during limb development. It controls outgrowth and patterning in the proximo-distal axis. In the last few years a considerable amount of new data regarding the cellular and molecular mechanisms underlying AER function and structure has been obtained. In this review, we describe and discuss current knowledge of the regulatory networks which control the induction, maturation and regression of the AER, as well as the link between Dorso-ventral patterning and the formation and position of the AER. Our aim is to integrate both recent and old knowledge to produce a wider picture of the AER which enhances our understanding of this relevant structure.

Arch Toxicol. 2009 Apr; 83(4): 319-33
Stapleton AR, Chan VT

Chlorpyrifos (CPF), a commonly used organophosphorus insecticide, induces acetylcholinesterase inhibition and cholinergic toxicity. Subtoxic exposure to CPF has long-term adverse effects on synaptic function/development and behavioral performance. To gain insight into the possible mechanism(s) of these observations, this study aims to investigate gene expression changes in the forebrain of rats treated with subtoxic CPF doses using DNA microarrays. Statistical analysis revealed that CPF treatment resulted in differential expression of 277 genes. Gene ontology and pathway analyses revealed that these genes have important roles in nervous system development and functions including axon guidance, Dorso-ventral axis formation, long-term potentiation, synaptic transmission, and insulin signaling. The results of biological associated network analysis showed that Gsk3b is highly connected in several of these networks suggesting its potential role in cellular response to CPF exposure/neurotoxicity. These findings might serve as the basis for future mechanistic analysis of the long-term adverse effects of subtoxic CPF exposure.

Zoolog Sci. 2000 Nov 1; 17(8): 1107-13
Onitsuka I, Takeda M, Maéno M

Msx is a homeodomain-containing transcriptional factor that plays an essential role in pattern formation in vertebrata and invertebrata embryos. In Xenopus laevis, two msx genes have been identified (Xmsx-1 and Xmsx-2). In the present study, we attempted to elucidate the expression and function of Xmsx-2B (formerly designated as Xhox7.1') in early embryogenesis. Whole mount in situ hybridization analyses showed that the expression pattern of Xmsx-2B at gastrula and neurula stages was very similar to that of Xmsx-1: the transcript of Xmsx-2B was observed in ventral and lateral sides of the embryo. At the tailbud stage, however, the expression pattern of Xmsx-2B in neural tissues was distinct from that of Xmsx-1. An RNA injection experiment revealed that, like BMP-4, Xmsx-2B has a strong ventralizing activity. In the Xmsx-2B -injected embryos, differentiation of axial structures such as the notochord, muscle, and neural tissue was completely suppressed, whereas alpha-globin mRNA, a blood cell marker, was highly expressed. Simultaneous injection of Xmsx-1 and Xmsx-2B RNAs showed that they function in an additive manner. It was also shown that coinjection of Xmsx-2B with a dominant-negative BMP-4 receptor (tBR), which can induce formation of secondary axis when injected alone in ventral blastomeres, suppressed secondary axis formation. Furthermore, Xmsx-2B also suppressed secondary axis formation, which was induced by a dominant-negative form of Xmsx-1 (VP16/msx-1). Therefore, like Xmsx-1, Xmsx-2B is a downstream nuclear factor of the BMP-4-derived ventralizing signal, and these two factors probably share the same target molecules. In conclusion, Xmsx-1 and Xmsx-2B function in Dorso-ventral axis formation in early Xenopus laevis development.

Gene Expr Patterns. 2008 Apr; 8(4): 217-26
Aquino JB, Marmigère F, Lallemend F, Lundgren TK, Villar MJ, Wegner M, Ernfors P

NEDD9 is a scaffolding protein in the integrin signaling pathway that is involved in cell adhesion dynamics. Little is known of the cellular localization of NEDD9 expression during embryonic development. In the present study, we have analyzed NEDD9 mRNA expression in the mouse and identified new relevant expression sites. In addition, we have characterized NEDD9 protein expression pattern for the first time in mammals. At E9.5-E10.5, high levels of Nedd9 and the neurogenic transcription factor neurogenin-2 (Ngn2) were found to largely overlap in two discrete domains of the trunk neural tube along its Dorso-ventral axis, with Nedd9 extending to more ventral regions of the ventricular zone and Ngn2 differentially expressed in neuronally committed progenitors of the intermediate zone. At encephalic and trunk levels of the neural tube, NEDD9 was present in Sox2(+) progenitor cell populations mostly generating Ngn2(+) and/or Nurr1(+) cells. A sharp down-regulation of NEDD9 expression was found in cells upon lineage commitment, as observed in Nurr1(+) and Ngn2(+) mesencephalic dopaminergic and brainstem neuronal progenitors. In other tissues/organs, i.e. prospective heart, retina, olfactory epithelium, gonads, cartilage, gut and pituitary gland, NEDD9 was found to be co-expressed with Sox2, RXR alpha and/or Nurr1-like proteins, suggesting that NEDD9 expression is confined to early progenitors involved in diverse organogenesis and that it may depend on the repertoire and levels of retinoic acid co-receptors expressed by those cells.

Mech Dev. 2008 Mar-Apr; 125(3-4): 353-72
Aw S, Adams DS, Qiu D, Levin M

Consistent laterality is a fascinating problem, and study of the Xenopus embryo has led to molecular characterization of extremely early steps in left-right patterning: bioelectrical signals produced by ion pumps functioning upstream of asymmetric gene expression. Here, we reveal a number of novel aspects of the H+/K+-ATPase module in chick and frog embryos. Maternal H+/K+-ATPase subunits are asymmetrically localized along the left-right, Dorso-ventral, and animal-vegetal axes during the first cleavage stages, in a process dependent on cytoskeletal organization. Using a reporter domain fused to molecular motors, we show that the cytoskeleton of the early frog embryo can provide asymmetric, directional information for subcellular transport along all three axes. Moreover, we show that the Kir4.1 potassium channel, while symmetrically expressed in a dynamic fashion during early cleavages, is required for normal LR asymmetry of frog embryos. Thus, Kir4.1 is an ideal candidate for the K+ ion exit path needed to allow the electroneutral H+/K+-ATPase to generate voltage gradients. In the chick embryo, we show that H+/K+-ATPase and Kir4.1 are expressed in the primitive streak, and that the known requirement for H+/K+-ATPase function in chick asymmetry does not function through effects on the circumferential expression pattern of Connexin43. These data provide details crucial for the mechanistic modeling of the physiological events linking subcellular processes to large-scale patterning and suggest a model where the early cytoskeleton sets up asymmetric ion flux along the left-right axis as a system of planar polarity functioning orthogonal to the apical-basal polarity of the early blastomeres.

Dev Biol. 2007 Dec 1; 312(1): 272-83
Murciano C, Pérez-Claros J, Smith A, Avaron F, Fernández TD, Durán I, Ruiz-Sánchez J, García F, Becerra J, Akimenko MA, Marí-Beffa M

The fins of actinopterygian can regenerate following amputation. Classical papers have shown that the ray, a structural unit of these fins, might regenerate independent of this appendage. Each fin ray is formed by two apposed contralateral hemirays. A hemiray may autonomously regenerate and segmentate in a position-independent manner. This is observed when heterotopically grafted into an interray space, after amputation following extirpation of the contralateral hemiray or when simply ablated. During this process, a proliferating hemiblastema is formed, as shown by bromodeoxyuridine incorporation, from which the complete structure will regenerate. This hemiblastema shows a patterning of gene expression domain similar to half ray blastema. Interactions between contralateral hemiblastema have been studied by recombinant rays composed of hemirays from different origins on the proximo-distal or Dorso-ventral axis of the caudal fin. Dye 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocianine perchlorate labeling of grafted tissues was used as tissular marker. Our results suggest both that there are contralateral interactions between hemiblastema of each ray, and that hemiblastema may vary its morphogenesis, always differentiating as their host region. These non-autonomous, position-dependent interactions control coordinated bifurcations, segment joints and ray length independently. A morphological study of the developing and regenerating fin of another long fin mutant zebrafish suggests that contralateral hemiblastema interactions are perturbed in this mutant.

Dev Dyn. 2007 Nov; 236(11): 2993-3006
Li N, Volff JN, Wizenmann A

The mouse Rab23 protein, a Ras-like GTPase, inhibits signaling through the Sonic hedgehog pathway and thus exerts a role in the dorsoventral patterning of the spinal cord. Rab23 mouse mutant embryos lack dorsal spinal cord cell types. We cloned the chicken Rab23 gene and studied its expression in the developing nervous system. Chick Rab23 mRNA is initially expressed in the entire neural tube but retracts to the dorsal alar plate. Unlike in mouse, we find Rab23 in chick already expressed asymmetrically during gastrulation. Ectopic expression of Rab23 in ventral midbrain induced dorsal genes (Pax3, Pax7) ectopically and reduced ventral genes (Nkx2.2 and Nkx6) without influencing cell proliferation or neurogenesis. Thus, in the developing brain of chick embryos Rab23 acts in the same manner as described for the caudal spinal cord in mouse. These data indicate that Rab23 plays an important role in patterning the Dorso-ventral axis by dorsalizing the neural tube.

J Cell Physiol. 2008 Feb; 214(2): 483-90
Soto X, Mayor R, Torrejón M, Montecino M, Hinrichs MV, Olate J

The non-canonical Wnt/Ca2+ signaling pathway has been implicated in the regulation of axis formation and gastrulation movements during early Xenopus laevis embryo development, by antagonizing the canonical Wnt/beta-catenin dorsalizing pathway and specifying ventral cell fate. However, the molecular mechanisms involved in this antagonist crosstalk are not known. Since Galphaq is the main regulator of Ca2+ signaling in vertebrates and from this perspective probably involved in the events elicited by the non-canonical Wnt/Ca2+ pathway, we decided to study the effect of wild-type Xenopus Gq (xGalphaq) in Dorso-ventral axis embryo patterning. Overexpression of xGalphaq or its endogenous activation at the dorsal animal region of Xenopus embryo both induced a strong ventralized phenotype and inhibited the expression of dorsal-specific mesoderm markers goosecoid and chordin. Dorsal expression of an xGalphaq dominant-negative mutant reverted the xGalphaq-induced ventralized phenotype. Finally, we observed that the Wnt8-induced secondary axis formation is reverted by endogenous xGalphaq activation, indicating that it is negatively regulating the Wnt/beta-catenin pathway.

PLoS One. 2007; 2(4): e374
Coolen M, Sauka-Spengler T, Nicolle D, Le-Mentec C, Lallemand Y, Da Silva C, Plouhinec JL, Robert B, Wincker P, Shi DL, Mazan S

The genetic mechanisms that control the establishment of early polarities and their link with embryonic axis specification and patterning seem to substantially diverge across vertebrates. In amphibians and teleosts, the establishment of an early Dorso-ventral polarity determines both the site of axis formation and its rostro-caudal orientation. In contrast, amniotes retain a considerable plasticity for their site of axis formation until blastula stages and rely on signals secreted by extraembryonic tissues, which have no clear equivalents in the former, for the establishment of their rostro-caudal pattern. The rationale for these differences remains unknown. Through detailed expression analyses of key development genes in a chondrichthyan, the dogfish Scyliorhinus canicula, we have reconstructed the ancestral pattern of axis specification in jawed vertebrates. We show that the dogfish displays compelling similarities with amniotes at blastula and early gastrula stages, including the presence of clear homologs of the hypoblast and extraembryonic ectoderm. In the ancestral state, these territories are specified at opposite poles of an early axis of bilateral symmetry, homologous to the Dorso-ventral axis of amphibians or teleosts, and aligned with the later forming embryonic axis, from head to tail. Comparisons with amniotes suggest that a dorsal expansion of extraembryonic ectoderm, resulting in an apparently radial symmetry at late blastula stages, has taken place in their lineage. The synthesis of these results with those of functional analyses in model organisms supports an evolutionary link between the Dorso-ventral polarity of amphibians and teleosts and the embryonic-extraembryonic organisation of amniotes. It leads to a general model of axis specification in gnathostomes, which provides a comparative framework for a reassessment of conservations both among vertebrates and with more distant metazoans.

Differentiation. 2007 Oct; 75(8): 726-36
Reza HM, Takahashi Y, Yasuda K

Dorso-ventral and proximo-distal axis formation of the optic cup is apparent from early stages of development. Pax6 is initially detectable in the optic vesicle and later shows a distal-high and proximal-low gradient of expression in the retina. To determine the early role of Pax6 in pattern formation of the optic cup, we expressed Pax6 ectopically in the optic vesicle of stages 9-10 chick embryos by in ovo electroporation, which resulted in a small eye-like phenotype. The signaling molecule fibroblast growth factor (FGF)8, which appears to be restricted to the central retina, was increased, whereas bone morphogenetic protein (BMP)4 and Tbx5, two dorsal markers, were down-regulated in Pax6-electroporated eye. Pax6 overexpression also decreased the expression of the ventral marker Vax. Electroporation with a dominant-negative form of Pax6 resulted in a decrease in FGF8 expression, but BMP4 expression was unaffected initially while it was diminished later. Our data suggest a new role for Pax6 in regulating FGF8 and BMP4 expression during pattern formation of the optic cup, and that a Pax6-regulated balance between FGF8 and BMP4 is critical for retinogenesis.

Gene Expr Patterns. 2007 Apr; 7(5): 584-95
Sprecher SG, Reichert H, Hartenstein V

The brain of Drosophila is formed by approximately 100 lineages, each lineage being derived from a stem cell-like neuroblast that segregates from the procephalic neurectoderm of the early embryo. A neuroblast map has been established in great detail for the early embryo, and a suite of molecular markers has been defined for all neuroblasts included in this map [Urbach, R., Technau, G.M. (2003a) Molecular markers for identified neuroblasts in the developing brain of Drosophila. Development 130, 3621-3637]. However, the expression of these markers was not followed into later embryonic or larval stages, mainly due to the fact that anatomical landmarks to which expression patterns could be related had not been defined. Such markers, in the form of stereotyped clusters of neurons whose axons project along cohesive bundles ("primary axon bundles" or "PABs") are now available [Younossi-Hartenstein, A., Nguyen, B., Shy, D., Hartenstein, V. 2006. Embryonic origin of the Drosophila brain neuropile. J. Comp. Neurol. 497, 981-998]. In the present study we have mapped the expression of molecular markers in relationship to primary neuronal clusters and their PABs. The markers we analyzed include many of the genes involved in patterning of the brain along the anteroposterior axis (cephalic gap genes, segment polarity genes) and Dorso-ventral axis (columnar patterning genes), as well as genes expressed in the dorsal protocerebrum and visual system (early eye genes). Our analysis represents an important step along the way to identify neuronal lineages of the mature brain with genes expressed in the early embryo in discrete neuroblasts. Furthermore, the analysis helped us to reconstruct the morphogenetic movements that transform the two-dimensional neuroblast layer of the early embryo into the three-dimensional larval brain and provides the basis for deeper understanding of how the embryonic brain develops.

Biochem Soc Symp. 2007; 9-22
Mikoshiba K

The IP3R [IP3 (inositol 1,4,5-trisphosphate) receptor] is responsible for Ca2+ release from the ER (endoplasmic reticulum). We have been working extensively on the P400 protein, which is deficient in Purkinje-neuron-degenerating mutant mice. We have discovered that P400 is an IP3R and we have determined the primary sequence. Purified IP3R, when incorporated into a lipid bilayer, works as a Ca2+ release channel and overexpression of IP3R shows enhanced IP3 binding and channel activity. Addition of an antibody blocks Ca2+ oscillations indicating that IP3R1 works as a Ca2+ oscillator. Studies on the role of IP3R during development show that IP3R is involved in fertilization and is essential for determination of Dorso-ventral axis formation. We found that IP3R is involved in neuronal plasticity. A double homozygous mutant of IP3R2 (IP3R type 2) and IP3R3 (IP3R type 3) shows a deficit of saliva secretion and gastric juice secretion suggesting that IP3Rs are essential for exocrine secretion. IP3R has various unique properties: cryo-EM (electron microscopy) studies show that IP3R contains multiple cavities; IP3R allosterically and dynamically changes its form reversibly (square form-windmill form); IP3R is functional even though it is fragmented by proteases into several pieces; the ER forms a meshwork but also forms vesicular ER and moves along microtubules using a kinesin motor; X ray analysis of the crystal structure of the IP3 binding core consists of an N-terminal beta-trefoil domain and a C-terminal alpha-helical domain. We have discovered ERp44 as a redox sensor in the ER which binds to the luminal part of IP3R1 and regulates its activity. We have also found the role of IP3 is not only to release Ca2+ but also to release IRBIT which binds to the IP3 binding core of IP3R.