Mouse Zic5 deficiency results in neural tube defects and hypoplasia of cephalic neural crest derivatives

Zic family genes encode zinc finger proteins, which are homologues of the Drosophila pair-rule gene odd-paired. In the present study, we characterized the fifth member of the mouse Zic family gene, mouse Zic5. Zic5 is located near Zic2, which is responsible for human brain malformation syndrome (holoprosencephaly, or HPE). In embryonic stages, Zic5 was expressed in dorsal part of neural tissues and limbs. Expression of Zic5 overlapped with those of other Zic genes, most closely with Zic2, but was not identical. Targeted disruption of Zic5 resulted in insufficient neural tube closure at the rostral end, similar to that seen in Zic2 mutant mice. In addition, the Zic5-deficient mice exhibited malformation of neural-crest-derived facial bones, especially the mandible, which had not been observed in other Zic family mutants. During the embryonic stages, there were delays in the development of the first branchial arch and extension of the trigeminal and facial nerves. Neural crest marker staining revealed fewer neural crest cells in the dorsal cephalic region of the mutant embryos without significant changes in their migration. When mouse Zic5 was overexpressed in Xenopus embryos, expression of a neural crest marker was enhanced. These findings suggested that Zic5 is involved in the generation of neural crest tissue in mouse development. ZIC5 is also located close to ZIC2 in humans, and deletions of 13q32, where ZIC2 is located, lead to congenital brain and digit malformations known as the "13q32 deletion syndrome". Based on both their similar expression pattern in mouse embryos and the malformations observed in Zic5-deficient mutant mice, human ZIC5 might be involved in the deletion syndrome.     

Expression of DDSG1, a novel gene encoding a putative DNA-binding protein in the embryonic chicken nervous system

In an attempt to clone genes expressed in the gizzard of the chicken embryo by differential display, we obtained a cDNA of a gene encoding a protein with a putative nuclear localization signal and a DNA-binding motif and designated it DDSG1 (differential display-screened gene expressed in the gizzard-1). Besides its expression in the gizzard, the gene is expressed in central and peripheral nervous systems such as brain, spinal cord and dorsal root ganglia in specific patterns.     

Multiple developmental programs are altered by loss of Zic1 and Zic4 to cause Dandy-Walker malformation cerebellar pathogenesis

Heterozygous deletions encompassing the ZIC1;ZIC4 locus have been identified in a subset of individuals with the common cerebellar birth defect Dandy-Walker malformation (DWM). Deletion of Zic1 and Zic4 in mice produces both cerebellar size and foliation defects similar to human DWM, confirming a requirement for these genes in cerebellar development and providing a model to delineate the developmental basis of this clinically important congenital malformation. Here, we show that reduced cerebellar size in Zic1 and Zic4 mutants results from decreased postnatal granule cell progenitor proliferation. Through genetic and molecular analyses, we show that Zic1 and Zic4 have Shh-dependent function promoting proliferation of granule cell progenitors. Expression of the Shh-downstream genes Ptch1, Gli1 and Mycn was downregulated in Zic1/4 mutants, although Shh production and Purkinje cell gene expression were normal. Reduction of Shh dose on the Zic1(+/-);Zic4(+/-) background also resulted in cerebellar size reductions and gene expression changes comparable with those observed in Zic1(-/-);Zic4(-/-) mice. Zic1 and Zic4 are additionally required to pattern anterior vermis foliation. Zic mutant folial patterning abnormalities correlated with disrupted cerebellar anlage gene expression and Purkinje cell topography during late embryonic stages; however, this phenotype was Shh independent. In Zic1(+/-);Zic4(+/-);Shh(+/-), we observed normal cerebellar anlage patterning and foliation. Furthermore, cerebellar patterning was normal in both Gli2-cko and Smo-cko mutant mice, where all Shh function was removed from the developing cerebellum. Thus, our data demonstrate that Zic1 and Zic4 have both Shh-dependent and -independent roles during cerebellar development and that multiple developmental disruptions underlie Zic1/4-related DWM.     

Overlapping expression pattern of the actin organizers Spir-1 and formin-2 in the developing mouse nervous system and the adult brain

The Wiskott-Aldrich homology domain 2 (WH2) family protein Spir and the formin Cappuccino belong to two distinct classes of actin organizers. Despite their functional classification as actin organizers, a major defect of Drosophila spire and cappuccino mutant oocytes is a failure in the orientation of microtubule plus ends towards the posterior pole. Mammalian homologues of spire are the spir-1 and spir-2 genes. The mouse and human formin-1 and formin-2 genes have high similarity to the cappuccino gene. The mouse formin-2 gene has been found to be expressed in the developing nervous system and in neuronal cells of the adult brain. By analyzing the expression of the spir-1 gene we show that spir-1 and formin-2 have a nearly identical expression pattern during mouse embryogenesis and in the adult brain. In mouse embryos both genes are expressed in the developing nervous system. In the adult brain high expression of the genes was found in the Purkinje cells of the cerebellum and in neuronal cells of the hippocampus and dentate gyrus.     

Cloning and characterization of the mouse Interleukin Enhancer Binding Factor 3 (Ilf3) homolog in a screen for RNA binding proteins

In a screen for RNA-binding proteins expressed during murine spermatogenesis, we have identified a cDNA that encodes a protein of 911 amino acids that contains two copies of the double-stranded RNA-binding motif and has 80% identity with human Interleukin Enhancer Binding Factor 3 (ILF3). Linkage and cytogenetic analyses localized the Ilf3 cDNA to a portion of mouse Chr 9, which shows conserved synteny with a region of human Chr 19 where the human ILF3 gene had been previously localized, supporting that we had cloned the murine homolog of ILF3. Northern analysis indicated the Ilf3 gene is ubiquitously expressed in mouse adult tissues with high levels of expression in the brain, thymus, testis, and ovary. Polyclonal antibodies detected multiple protein species in a subset of the tissues expressing Ilf3 RNA. Immunoreactive species are present at high levels in the thymus, testis, ovary, and the spleen to a lesser extent. The high degree of sequence similarity between the mouse ILF3 protein and other dsRNA binding motif-containing proteins suggests a role in RNA metabolism, while the differential expression indicates the mouse ILF3 protein predominantly functions in tissues containing developing lymphocyte and germ cells. Received: 21 October 1998 / Accepted: 15 January 1999     

Expression of Plxdc2/TEM7R in the developing nervous system of the mouse

Plexin-domain containing 2 (Plxdc2) is a relatively uncharacterised transmembrane protein with an area of nidogen homology and a plexin repeat (PSI domain) in its extracellular region. Here, we describe Plxdc2 expression in the embryonic mouse, with particular emphasis on the developing central nervous system. Using light microscopy and optical projection tomography (OPT), we analyse RNA in situ hybridization patterns and expression of two reporter genes, beta-geo (a fusion of beta-galactosidase to neomycin phosphotransferase) and placental alkaline phosphatase (PLAP) in a Plxdc2 gene trap mouse line (KST37; [Leighton, P.A., Mitchell, K.J., Goodrich, L.V., Lu, X., Pinson, K., Scherz, P., Skarnes, W.C., Tessier-Lavigne, M., 2001. Defining brain wiring patterns and mechanisms through gene trapping in mice. Nature 410, 174-179]). At mid-embryonic stages (E9.5-E11.5) Plxdc2-betageo expression is prominent in a number of patterning centres of the brain, including the cortical hem, midbrain-hindbrain boundary and the midbrain floorplate. Plxdc2 is expressed in other tissues, most notably the limbs, lung buds and developing heart, as well as the spinal cord and dorsal root ganglia. At E15.5, expression is apparent in a large number of discrete nuclei and structures throughout the brain, including the glial wedge and derivatives of the cortical hem. Plxdc2-betageo expression is particularly strong in the developing Purkinje cell layer, especially in the posterior half of the cerebellum. The PLAP marker is expressed in a number of axonal tracts, including the posterior commissure, mammillotegmental tract and cerebellar peduncle. We compare Plxdc2-betageo expression in the embryonic brain with the much more restricted expression of the related gene Plxdc1 and with members of the Wnt family (Wnt3a, Wnt5a and Wnt8b) that show a striking overlap with Plxdc2 expression in certain areas.