Christoph Wiegreffe

A dual function of Bcl11b/Ctip2 in hippocampal neurogenesis

The development of the dentate gyrus is characterized by distinct phases establishing a durable stem‐cell pool required for postnatal and adult neurogenesis. Here, we report that Bcl11b/Ctip2, a zinc finger transcription factor expressed in postmitotic neurons, plays a critical role during postnatal development of the dentate gyrus. Forebrain‐specific ablation of Bcl11b uncovers dual phase‐specific functions of Bcl11b demonstrated by feedback control of the progenitor cell compartment as well as regulation of granule cell differentiation, leading to impaired spatial learning and memory in mutants. Surprisingly, we identified Desmoplakin as a direct transcriptional target of Bcl11b. Similarly to Bcl11b, postnatal neurogenesis and granule cell differentiation are impaired in Desmoplakin mutants. Re‐expression of Desmoplakin in Bcl11b mutants rescues impaired neurogenesis, suggesting Desmoplakin to be an essential downstream effector of Bcl11b in hippocampal development. Together, our data define an important novel regulatory pathway in hippocampal development, by linking transcriptional functions of Bcl11b to Desmoplakin, a molecule known to act on cell adhesion.

Sclerotomal origin of smooth muscle cells in the wall of the avian dorsal aorta

The dorsal aorta is the earliest formed intraembryonic blood vessel. It is composed of an inner lining consisting of endothelial cells and an outer wall consisting of smooth muscle cells (SMCs) and fibrocytes. Aortic SMCs have been suggested to arise from several developmental lineages. Cephalic neural crest provides SMCs of the proximal part of the aorta, and SMCs of the distal part are derived from the paraxial mesoderm. Here, we show by using quail‐chick chimerization that in the avian embryo, SMCs in the wall of the dorsal aorta at trunk level arise from the sclerotome. Our findings indicate a two‐step process of aortic wall formation. First, non‐paraxial mesoderm‐derived mural cells accumulate at the floor of the aorta. We refer to these cells as primary SMCs. Second, SMCs from the sclerotome are recruited to the roof and sides of the aorta, eventually replacing the primary SMCs in the aortic floor. Developmental Dynamics 236:2578–2585, 2007.

Bcl11a is required for neuronal morphogenesis and sensory circuit formation in dorsal spinal cord development

Dorsal spinal cord neurons receive and integrate somatosensory information provided by neurons located in dorsal root ganglia. Here we demonstrate that dorsal spinal neurons require the Krüppel-C2H2 zinc-finger transcription factor Bcl11a for terminal differentiation and morphogenesis. The disrupted differentiation of dorsal spinal neurons observed in Bcl11a mutant mice interferes with their correct innervation by cutaneous sensory neurons. To understand the mechanism underlying the innervation deficit, we characterized changes in gene expression in the dorsal horn of Bcl11a mutants and identified dysregulated expression of the gene encoding secreted frizzled-related protein 3 (sFRP3, or Frzb). Frzb mutant mice show a deficit in the innervation of the spinal cord, suggesting that the dysregulated expression of Frzb can account in part for the phenotype of Bcl11a mutants. Thus, our genetic analysis of Bcl11a reveals essential functions of this transcription factor in neuronal morphogenesis and sensory wiring of the dorsal spinal cord and identifies Frzb, a component of the Wnt pathway, as a downstream acting molecule involved in this process.

Bcl11a (Ctip1) Controls Migration of Cortical Projection Neurons through Regulation of Sema3c

During neocortical development, neurons undergo polarization, oriented migration, and layer-type-specific differentiation. The transcriptional programs underlying these processes are not completely understood. Here, we show that the transcription factor Bcl11a regulates polarity and migration of upper layer neurons. Bcl11a-deficient late-born neurons fail to correctly switch from multipolar to bipolar morphology, resulting in impaired radial migration. We show that the expression of Sema3c is increased in migrating Bcl11a-deficient neurons and that Bcl11a is a direct negative regulator of Sema3c transcription. In vivo gain-of-function and rescue experiments demonstrate that Sema3c is a major downstream effector of Bcl11a required for the cell polarity switch and for the migration of upper layer neurons. Our data uncover a novel Bcl11a/Sema3c-dependent regulatory pathway used by migrating cortical neurons.

Communication between distant epithelial cells by filopodia-like protrusions during embryonic development

Long-range intercellular communication is essential for the regulation of embryonic development. Apart from simple diffusion, various modes of signal transfer have been described in the literature. Here, we describe a novel type of cellular extensions found in epithelial cells of the somites in chicken embryos. These filopodia-like protrusions span the subectodermal space overlying the dorsal surface of the somites and contact the ectoderm. We show that these protrusions are actin- and tubulin-positive and require Rac1 for their formation. The presence of glycophosphatidylinositol-anchored proteins and net retrograde trafficking of the transmembrane Wnt-receptor Frizzled-7 along the protrusions indicate their role in signal transport and distribution. Taken together, our data suggest a role of filopodia-like protrusions in mediating signaling events between distant epithelial cells during embryonic development. Summary: Epithelial cells of chick embryo somites exhibit filopodia-like protrusions that serve as transport conduits connecting them to the overlying surface ectoderm.

Remodeling of aortic smooth muscle during avian embryonic development

The dorsal aorta is the earliest formed intraembryonic blood vessel in vertebrates composed of an inner lining of endothelial cells (ECs) and a slightly later‐forming outer wall consisting of vascular smooth muscle cells (SMCs) and pericytes. We previously identified the sclerotome as the only somitic compartment contributing to aortic SMCs in the trunk of the avian embryo. However, we demonstrated that the first SMCs in the aortic floor are not of somitic origin and must be derived from a different source. Here, we show that the primary SMCs are a transient population of aortic wall cells originating from the splanchnic mesoderm. A model is presented suggesting that wall formation of the early dorsal aorta in chick is a two‐step process: The primary, transient SMCs in the aortic floor originate in the splanchnic mesoderm, whereas the secondary, definitive SMCs of the entire aortic wall originate in the sclerotome. Developmental Dynamics 238:624–631, 2009.

Somite compartments in anamniotes

Somites are a common feature of the phylotypic stage of embryos of all higher chordates. In amniote species like mouse and chick, somite development has been the subject of intense research over many decades, giving insight into the morphological and molecular processes leading to somite compartmentalization and subsequent differentiation. In anamniotes, somite development is much less understood. Except for recent data from zebrafish, and morphological studies in Xenopus, very little is known about the formation of somite compartments and the differentiation of somite derivatives in anamniotes. Here, we give a brief overview on the development of myotome, sclerotome and dermomyotome in various anamniote organisms, and point out the different mechanisms of somite development between anamniotes and the established amniote model systems.

Integration of human model neurons (NT2) into embryonic chick nervous system

Postmitotic neurons were generated from the human NT2 teratocarcinoma cell line in a novel cell aggregate differentiation procedure. Approximately a third of the differentiated neurons expressed cell markers related to cholinergic neurotransmission. To examine whether this human cell model system can be directed toward a motoneuronal fate, postmitotic neurons were co‐cultured with mouse myotubes. Outgrowing neuronal processes established close contact with the myotubes and formed neuromuscular junction‐like structures that bound α‐bungarotoxin. To determine how grafted precursor cells and neurons respond to embryonic nerve tissue, NT2 cells at different stages of neural development were injected into chick embryo neural tube and brain. Grafted NT2 neurons populated both parts of the nervous system, sometimes migrating away from the site of injection. The neural tube appeared to be more permissive for neurite extensions than the brain. Moreover, extending neurites of spinal grafts were approaching the ventral roots, thus resembling motoneuronal projections. Developmental Dynamics 239:496–504, 2010.

Thirty-eight-negative kinase 1 mediates trauma-induced intestinal injury and multi-organ failure

Dysregulated intestinal epithelial apoptosis initiates gut injury, alters the intestinal barrier, and can facilitate bacterial translocation leading to a systemic inflammatory response syndrome (SIRS) and/or multi-organ dysfunction syndrome (MODS). A variety of gastrointestinal disorders, including inflammatory bowel disease, have been linked to intestinal apoptosis. Similarly, intestinal hyperpermeability and gut failure occur in critically ill patients, putting the gut at the center of SIRS pathology. Regulation of apoptosis and immune-modulatory functions have been ascribed to Thirty-eight-negative kinase 1 (TNK1), whose activity is regulated merely by expression. We investigated the effect of TNK1 on intestinal integrity and its role in MODS. TNK1 expression induced crypt-specific apoptosis, leading to bacterial translocation, subsequent septic shock, and early death. Mechanistically, TNK1 expression in vivo resulted in STAT3 phosphorylation, nuclear translocation of p65, and release of IL-6 and TNF-&agr;. A TNF-&agr; neutralizing antibody partially blocked development of intestinal damage. Conversely, gut-specific deletion of TNK1 protected the intestinal mucosa from experimental colitis and prevented cytokine release in the gut. Finally, TNK1 was found to be deregulated in the gut in murine and porcine trauma models and human inflammatory bowel disease. Thus, TNK1 might be a target during MODS to prevent damage in several organs, notably the gut.

Time-lapse Confocal Imaging of Migrating Neurons in Organotypic Slice Culture of Embryonic Mouse Brain Using In Utero Electroporation

In utero electroporation is a rapid and powerful approach to study the process of radial migration in the cerebral cortex of developing mouse embryos. It has helped to describe the different steps of radial migration and characterize the molecular mechanisms controlling this process. To directly and dynamically analyze migrating neurons they have to be traced over time. This protocol describes a workflow that combines in utero electroporation with organotypic slice culture and time-lapse confocal imaging, which allows for a direct examination and dynamic analysis of radially migrating cortical neurons. Furthermore, detailed characterization of migrating neurons, such as migration speed, speed profiles, as well as radial orientation changes, is possible. The method can easily be adapted to perform functional analyses of genes of interest in radially migrating cortical neurons by loss and gain of function as well as rescue experiments. Time-lapse imaging of migrating neurons is a state-of-the-art technique that once established is a potent tool to study the development of the cerebral cortex in mouse models of neuronal migration disorders.