Heiner Westphal

Targeted disruption of the alpha isoform of the peroxisome proliferator-activated receptor gene in mice results in abolishment of the pleiotropic effects of peroxisome proliferators.

To gain insight into the function of peroxisome proliferator-activated receptor (PPAR) isoforms in rodents, we disrupted the ligand-binding domain of the alpha isoform of mouse PPAR (mPPAR alpha) by homologous recombination. Mice homozygous for the mutation lack expression of mPPAR alpha protein and yet are viable and fertile and exhibit no detectable gross phenotypic defects. Remarkably, these animals do not display the peroxisome proliferator pleiotropic response when challenged with the classical peroxisome proliferators, clofibrate and Wy-14,643. Following exposure to these chemicals, hepatomegaly, peroxisome proliferation, and transcriptional-activation of target genes were not observed. These results clearly demonstrate that mPPAR alpha is the major isoform required for mediating the pleiotropic response resulting from the actions of peroxisome proliferators. mPPAR alpha-deficient animals should prove useful to further investigate the role of this receptor in hepatocarcinogenesis, fatty acid metabolism, and cell cycle regulation.

The LIM‐only protein Lmo2 is a bridging molecule assembling an erythroid, DNA‐binding complex which includes the TAL1, E47, GATA‐1 and Ldb1/NLI proteins

The LIM‐only protein Lmo2, activated by chromosomal translocations in T‐cell leukaemias, is normally expressed in haematopoiesis. It interacts with TAL1 and GATA‐1 proteins, but the function of the interaction is unexplained. We now show that in erythroid cells Lmo2 forms a novel DNA‐binding complex, with GATA‐1, TAL1 and E2A, and the recently identified LIM‐binding protein Ldb1/NLI. This oligomeric complex binds to a unique, bipartite DNA motif comprising an E‐box, CAGGTG, followed ∼9 bp downstream by a GATA site. In vivo assembly of the DNA‐binding complex requires interaction of all five proteins and establishes a transcriptional transactivating complex. These data demonstrate one function for the LIM‐binding protein Ldb1 and establish a function for the LIM‐only protein Lmo2 as an obligatory component of an oligomeric, DNA‐binding complex which may play a role in haematopoiesis.

Sonic hedgehog is essential to foregut development

Congenital malformation of the foregut is common in humans, with an estimated incidence of 1 in 3000 live births, although its aetiology remains largely unknown. Mice with a targeted deletion of Sonic hedgehog ( Shh) have foregut defects that are apparent as early as embryonic day 9.5, when the tracheal diverticulum begins to outgrow. Homozygous Shh-null mutant mice show oesophageal atresia/stenosis, tracheo-oesophageal fistula and tracheal and lung anomalies, features similar to those observed in humans with foregut defects. The lung mesenchyme shows enhanced cell death, decreased cell proliferation and downregulation of Shh target genes. These results indicate that Shh is required for the growth and differentiation of the oesophagus, trachea and lung, and suggest that mutations in SHH and its signalling components may be involved in foregut defects in humans.

Dickkopf1 Is Required for Embryonic Head Induction and Limb Morphogenesis in the Mouse

Dickkopf1 (Dkk1) is a secreted protein that acts as a Wnt inhibitor and, together with BMP inhibitors, is able to induce the formation of ectopic heads in Xenopus. Here, we show that Dkk1 null mutant embryos lack head structures anterior of the midbrain. Analysis of chimeric embryos implicates the requirement of Dkk1 in anterior axial mesendoderm but not in anterior visceral endoderm for head induction. In addition, mutant embryos show duplications and fusions of limb digits. Characterization of the limb phenotype strongly suggests a role for Dkk1 both in cell proliferation and in programmed cell death. Our data provide direct genetic evidence for the requirement of secreted Wnt antagonists during embryonic patterning and implicate Dkk1 as an essential inducer during anterior specification as well as a regulator during distal limb patterning.

Specification of Pituitary Cell Lineages by the LIM Homeobox Gene Lhx3

During pituitary organogenesis, the progressive differentiation of distinct pituitary-specific cell lineages from a common primordium involves a series of developmental decisions and inductive interactions. Targeted gene disruption in mice showed that Lhx3, a LIM homeobox gene expressed in the pituitary throughout development, is essential for differentiation and proliferation of pituitary cell lineages. In mice homozygous for the Lhx3 mutation, Rathkes pouch formed but failed to grow and differentiate; such mice lacked both the anterior and intermediate lobes of the pituitary. The determination of all pituitary cell lineages, except the corticotrophs, was affected, suggesting that a distinct, Lhx3-independent ontogenetic pathway exists for the initial specification of this lineage.

Functions of LIM-homeobox genes.

Homeobox genes play fundamental roles in development. They can be subdivided into several subfamilies, one of which is the LIM-homeobox subfamily. The primary structure of LIM-homeobox genes has been remarkably conserved through evolution. Have their functions similarly been conserved? A host of new data has been derived from mutational analysis in diverse organisms, such as nematodes, flies and vertebrates. These studies have revealed a prominent involvement of LIM-homeodomain proteins in tissue patterning and differentiation, and their function in neural patterning is evident in all organisms studied to date. Here, we summarize the recent findings on LIM-homeobox gene function, compare the function of these genes from different organisms and describe specific co-factor requirements.

LIM Homeodomain Factors Lhx3 and Lhx4 Assign Subtype Identities for Motor Neurons

The circuits that control movement are comprised of discrete subtypes of motor neurons. How motor neuron subclasses develop and extend axons to their correct targets is still poorly understood. We show that LIM homeodomain factors Lhx3 and Lhx4 are expressed transiently in motor neurons whose axons emerge ventrally from the neural tube (v-MN). Motor neurons develop in embryos deficient in both Lhx3 and Lhx4, but v-MN cells switch their subclass identity to become motor neurons that extend axons dorsally from the neural tube (d-MN). Conversely, the misexpression of Lhx3 in dorsal-exiting motor neurons is sufficient to reorient their axonal projections ventrally. Thus, Lhx3 and Lhx4 act in a binary fashion during a brief period in development to specify the trajectory of motor axons from the neural tube.

Nitric oxide negatively regulates mammalian adult neurogenesis

Neural progenitor cells are widespread throughout the adult central nervous system but only give rise to neurons in specific loci. Negative regulators of neurogenesis have therefore been postulated, but none have yet been identified as subserving a significant role in the adult brain. Here we report that nitric oxide (NO) acts as an important negative regulator of cell proliferation in the adult mammalian brain. We used two independent approaches to examine the function of NO in adult neurogenesis. In a pharmacological approach, we suppressed NO production in the rat brain by intraventricular infusion of an NO synthase inhibitor. In a genetic approach, we generated a null mutant neuronal NO synthase knockout mouse line by targeting the exon encoding active center of the enzyme. In both models, the number of new cells generated in neurogenic areas of the adult brain, the olfactory subependyma and the dentate gyrus, was strongly augmented, which indicates that division of neural stem cells in the adult brain is controlled by NO and suggests a strategy for enhancing neurogenesis in the adult central nervous system.

The LIM homeobox gene Lhx9 is essential for mouse gonad formation.

During mammalian embryonic development, the ovaries and testes develop from somatic cells of the urogenital ridges as indifferent gonads, harbouring primordial germ cells that have migrated there. After sex determination of the gonads, the testes produce testosterone and anti-Mullerian hormone which mediate male sexual differentiation, and the female developmental pathway ensues in their absence. Here we show that transcripts of the LIM homeobox gene Lhx9 are present in urogenital ridges of mice at embryonic day 9.5; later they localize to the interstitial region as morphological differentiation occurs. In mice lacking Lhx9 function, germ cells migrate normally, but somatic cells of the genital ridge fail to proliferate and a discrete gonad fails to form. In the absence of testosterone and anti-Mullerian hormone, genetically male mice are phenotypically female. The expression of steroidogenic factor 1 (Sf1), a nuclear receptor essential for gonadogenesis, is reduced to minimal levels in the Lhx9-deficient genital ridge, indicating that Lhx9 may lie upstream of Sf1 in a developmental cascade. Unlike mice lacking other genes that mediate early stages of gonadogenesis, Lhx9 mutants do not exhibit additional major developmental defects. Thus, LHX9 mutations may underlie certain forms of isolated gonadal agenesis in humans.

Neurturin and glial cell line-derived neurotrophic factor receptor-beta (GDNFR-beta), novel proteins related to GDNF and GDNFR-alpha with specific cellular patterns of expression suggesting roles in the developing and adult nervous system and in peripheral organs.

Cloning strategies were used to identify a gene termed glial cell line-derived neurotrophic factor receptor-β (GDNFR-β) related to GDNFR-α. In situ hybridization was then used to map cellular expression of the GDNF-related trophic factor neurturin (NTN) and GDNFR-β mRNA in developing and adult mice, and comparisons with GDNFR-α and RET were made. Neurturin is expressed in postnatal cerebral cortex, striatum, several brainstem areas, and the pineal gland. GDNFR-β mRNA was more widely expressed in the developing and adult CNS, including cerebral cortex, cerebellum, thalamus, zona incerta, hypothalamus, brainstem, and spinal cord, and in subpopulations of sensory neurons and developing peripheral nerves. NTN colocalized with RET and GDNFR-α in ureteric buds of the developing kidney. The circular muscle layer of the developing intestines, smooth muscle of the urether, and developing bronchiolae also expressed NTN. GDNFR-β was found in myenteric but not submucosal intestinal plexuses. In developing salivary glands NTN had an epithelial expression, whereas GDNFR-β was expressed in surrounding tissue. Neurturin and GDNFR-β were present in developing sensory organs. In the gonads, NTN appeared to be expressed in Sertoli cells and in the epithelium of the oviduct, whereas GDNFR-β was expressed by the germ cell line. Our findings suggest multiple roles for NTN and GDNFR-β in the developing and adult organism. Although NTN and GDNFR-β expression patterns are sometimes complementary, this is not always the case, suggesting multiple modi operandi of GDNF and NTN in relation to RET and the two binding proteins, GDNFR-α and GDNFR-β.