Harold Cremer

The homeobox gene Phox2b is essential for the development of autonomic neural crest derivatives.

The sympathetic, parasympathetic and enteric ganglia are the main components of the peripheral autonomic nervous system, and are all derived from the neural crest. The factors needed for these structures to develop include the transcription factor Mash1 (refs 3,4,5), the glial-derived neurotrophic factor GNDF (refs 6,7,8) and its receptor subunits, and the neuregulin signalling system, each of which is essential for the differentiation and survival of subsets of autonomic neurons. Here we show that all autonomic ganglia fail to form properly and degenerate in mice lacking the homeodomain transcription factor Phox2b, as do the three cranial sensory ganglia that are part of the autonomic reflex circuits. In the anlagen of the enteric nervous system and the sympathetic ganglia, Phox2b is needed for the expression of the GDNF-receptor subunit Ret and for maintaining Mash1 expression. Mutant ganglionic anlagen also fail to switch on the genes that encode two enzymes needed for the biosynthesis of the neurotransmitter noradrenaline, dopamine-β-hydroxylase and tyrosine hydroxylase, demonstrating that Phox2b regulates the noradrenergic phenotype in vertebrates.

PSA–NCAM Is Required for Activity-Induced Synaptic Plasticity

Hippocampal organotypic slice cultures maintained 10-20 days in vitro express a high level of the polysialylated embryonic form of neural cell adhesion molecule (NCAM) (PSA-NCAM). Treatment of the cultures with endoneuraminidase-N selectively removed polysialic acid (PSA) from NCAM and completely prevented induction of long-term potentiation (LTP) and long-term depression (LTD) without affecting cellular or synaptic parameters. Similarly, slices prepared from transgenic mice lacking the NCAM gene exhibited a decaying LTP. No inhibition of N-methyl-D-aspartic acid receptor-dependent synaptic responses was detected. Washout of the enzyme resulted in reexpression of PSA immunoreactivity which correlated with a complete recovery of LTP and LTD. This reexpression was blocked by TTX and low calcium and enhanced by bicuculline. Taken together, these results indicate that neuronal activity regulates the expression of PSA-NCAM at the synapse and that this expression is required for the induction of synaptic plasticity.

Defects in sensory and autonomic ganglia and absence of locus coeruleus in mice deficient for the homeobox gene Phox2a

Phox2a is a vertebrate homeodomain protein expressed in subsets of differentiating neurons. Here, we show that it is essential for proper development of the locus coeruleus, a subset of sympathetic and parasympathetic ganglia and the VIIth, IXth, and Xth cranial sensory ganglia. In the sensory ganglia, we have identified two differentiation blocks in Phox2a-/- mice. First, the transient expression of dopamine-beta-hydroxylase in neuroblasts is abolished, providing evidence that Phox2a controls noradrenergic traits in vivo. Second, the expression of the GDNF receptor subunit Ret is dramatically reduced, and there is a massive increase in apoptosis of ganglion cells, which are known to depend on GDNF in vivo. Therefore, Phox2a appears to regulate conventional differentiation traits and the ability of neurons to respond to essential survival factors.

Reelin is a detachment signal in tangential chain-migration during postnatal neurogenesis

During development, Reelin acts on migrating neuronal precursors and controls correct cell positioning in the cortex and other brain structures by a hitherto unidentified mechanism. Here we show that in the postnatal mouse brain, Reelin acts as a detachment signal for chain-migrating interneuron precursors in the olfactory bulb. Neuronal precursors cultured in Matrigel detached from chains and migrated individually in the presence of exogenously added Reelin protein or Reelin-expressing brain tissues. Furthermore, we found that in reeler mutant mice, neuronal precursors accumulated in the olfactory bulb and remained in clusters, indicating that they did not change from tangential chain-migration to radial individual migration. Our data provide direct evidence that Reelin acts as a detachment signal, but not a stop or guidance cue. We propose that Reelin may have comparable functions during development.

NCAM is essential for axonal growth and fasciculation in the hippocampus.

The neural cell adhesion molecule (NCAM), probably the best characterized and most abundant cell adhesion molecule on neurons, is thought to be a major regulator of axonal growth and pathfinding. Here we present a detailed analysis of these processes in mice deficient for all NCAM isoforms, generated by gene targeting. The hippocampal mossy fiber tract shows prominent expression of polysialylated NCAM and the generation of new axonal projections throughout life. Focusing on this important intrahippocampal connection, we demonstrate that in the absence of NCAM, fasciculation and pathfinding of these axons are strongly affected. In addition we show alterations in the distribution of mossy fiber terminals. The phenotype is more severe in adult than in young animals, suggesting an essential role for NCAM in the maintenance of plasticity in the mature nervous system.

Coupling between hydrodynamic forces and planar cell polarity orients mammalian motile cilia

In mammals, motile cilia cover many organs, such as fallopian tubes, respiratory tracts and brain ventricles. The development and function of these organs critically depend on efficient directional fluid flow ensured by the alignment of ciliary beating. To identify the mechanisms involved in this process, we analysed motile cilia of mouse brain ventricles, using biophysical and molecular approaches. Our results highlight an original orientation mechanism for ependymal cilia whereby basal bodies first dock apically with random orientations, and then reorient in a common direction through a coupling between hydrodynamic forces and the planar cell polarity (PCP) protein Vangl2, within a limited time-frame. This identifies a direct link between external hydrodynamic cues and intracellular PCP signalling. Our findings extend known PCP mechanisms by integrating hydrodynamic forces as long-range polarity signals, argue for a possible sensory role of ependymal cilia, and will be of interest for the study of fluid flow-mediated morphogenesis.

Combined Natural Killer Cell and Dendritic Cell Functional Deficiency in KARAP/DAP12 Loss-of-Function Mutant Mice

KARAP/DAP12 is a transmembrane polypeptide with an intracytoplasmic immunoreceptor tyrosine-based activation motif (ITAM). KARAP/DAP12 is associated with several activating cell surface receptors in hematopoietic cells. Here, we report that knockin mice bearing a nonfunctional KARAP/DAP12 ITAM present altered innate immune responses. Although in these mice NK cells are present and their repertoire of inhibitory MHC class I receptors is intact, the NK cell spectrum of natural cytotoxicity toward tumor cell targets is restricted. KARAP/DAP12 loss-of-function mutant mice also exhibit a dramatic accumulation of dendritic cells in muco-cutaneous epithelia, associated with an impaired hapten-specific contact sensitivity. Thus, despite its homology with CD3zeta and FcRgamma, KARAP/DAP12 plays a specific role in innate immunity, emphasizing the nonredundancy of these ITAM-bearing polypeptides in hematopoietic cells.

Molecular Interaction between Projection Neuron Precursors and Invading Interneurons via Stromal-Derived Factor 1 (CXCL12)/CXCR4 Signaling in the Cortical Subventricular Zone/Intermediate Zone

Most cortical interneurons are generated in the subpallial ganglionic eminences and migrate tangentially to their final destinations in the neocortex. Within the cortex, interneurons follow mainly stereotype routes in the subventricular zone/intermediate zone (SVZ/IZ) and in the marginal zone. It has been suggested that interactions between invading interneurons and locally generated projection neurons are implicated in the temporal and spatial regulation of the invasion process. However, so far experimental evidence for such interactions is lacking. We show here that the chemokine stromal-derived factor 1 (SDF-1; CXCL12) is expressed in the main invasion route for cortical interneurons in the SVZ/IZ. Most SDF-1-positive cells are proliferating and express the homeodomain transcription factors Cux1 and Cux2. Using MASH-1 mutant mice in concert with the interneuron marker DLX, we exclude that interneurons themselves produce the chemokine in an autocrine manner. We conclude that the SDF-1-expressing cell population represents the precursors of projection neurons during their transition and amplification in the SVZ/IZ. Using mice lacking the SDF-1 receptor CXCR4 or Pax6, we demonstrate that SDF-1 expression in the cortical SVZ/IZ is essential for recognition of this pathway by interneurons. These results represent the first evidence for a molecular interaction between precursors of projection neurons and invading interneurons during corticogenesis.

REDUCED EXPRESSION OF NEURAL CELL ADHESION MOLECULE INDUCES METASTATIC DISSEMINATION OF PANCREATIC BETA TUMOR CELLS

As in the development of many human cancers, in a transgenic mouse model of β-cell carcinogenesis (Rip1Tag2), expression of neural cell adhesion molecule (NCAM) changes from the 120-kDa isoform in normal tissue to the 140/180-kDa isoforms in tumors. NCAM-deficient Rip1Tag2 mice, generated by crossing Rip1Tag2 mice with NCAM knockout mice, develop metastases, a tumor stage that is not seen in normal Rip1Tag2 mice. In contrast, overexpression of NCAM 120 in NCAM-deficient Rip1Tag2 mice prevents tumor metastasis. The results indicate that the loss of NCAM-mediated cell adhesion is one rate-limiting step in the actual metastatic dissemination of β tumor cells.

Revisiting the function of PSA-NCAM in the nervous system.

Since its first description the polysialylated form of NCAM (PSA-NCAM) is thought to be a major regulator of cell-cell interactions in the nervous system. Over the past few years many crucial questions have been answered concerning PSA biosynthesis and function. Among these are the identification and cloning of the key enzymes that are responsible for its synthesis and the fact that expression of PSA is not restricted to developmental stages but maintained in the adult nervous system. In the adult, PSA has been shown to be not only a marker of structural plasticity but seems to be a major player in these processes. Originally suggested to be a purely anti-adhesive factor, modulating cell-cell interactions in general and by this allowing plasticity, there is now increasing evidence that this might not be the whole story. Instead, it appears possible that PSA-NCAM interacts with secreted signaling molecules and by this fulfills a more instructive function in brain plasticity.