Neural progenitor cells in the developing dorsal forebrain give rise to excitatory neurons, astrocytes, and oligodendrocytes for the neocortex. While we are starting to gain a better understanding about the mechanisms that direct the formation of neocortical neurons and astrocytes, far less is known about the molecular mechanisms that instruct dorsal forebrain progenitors to make oligodendrocytes. In this study, we show that Sonic hedgehog (Shh) signaling is required in dorsal progenitors for their late embryonic transition to oligodendrogenesis. Using genetic lineage-tracing in mice of both sexes, we demonstrate that most oligodendrocytes in the embryonic neocortex derive from Emx1 dorsal forebrain progenitors. Deletion of the Shh signaling effector specifically in Emx1 progenitors led to significantly decreased oligodendrocyte numbers in the embryonic neocortex. Conversely, knock-out of the Shh antagonist was sufficient to increase neocortical oligodendrogenesis. Using conditional knock-out strategies, we found that Shh ligand is supplied to dorsal progenitors through multiple sources. Loss of from Dlx5/6 interneurons caused a significant reduction in oligodendrocytes in the embryonic neocortex. This phenotype was identical to that observed upon deletion from the entire CNS using, indicating that interneurons migrating into the neocortex from the subpallium are the primary neural source of Shh for dorsal oligodendrogenesis. Additionally, deletion of from migrating interneurons together with the choroid plexus epithelium led to a more severe loss of oligodendrocytes, suggesting that the choroid plexus is an important non-neural source of Shh ligand. Together, our studies demonstrate that the dorsal wave of neocortical oligodendrogenesis occurs earlier than previously appreciated and requires highly regulated Shh signaling from multiple embryonic sources. Most neocortical oligodendrocytes are made by neural progenitors in the dorsal forebrain, but the mechanisms that specify this fate are poorly understood. This study identifies Sonic hedgehog (Shh) signaling as a critical pathway in the transition from neurogenesis to oligodendrogenesis in dorsal forebrain progenitors during late embryonic development. The timing of this neuron-to-glia "switch" coincides with the arrival of migrating interneurons into the dorsal germinal zone, which we identify as a critical source of Shh ligand, which drives oligodendrogenesis. Our data provide evidence for a new model in which Shh signaling increases in the dorsal forebrain late in embryonic development to provide a temporally regulated mechanism that initiates the third wave of neocortical oligodendrogenesis.

ABSTRACTSalmonella entericaserovar Typhimurium is a Gram-negative member of the familyEnterobacteriaceaeand is a common cause of bacterial food poisoning in humans. The fimbrial appendages are found on the surface of many enteric bacteria and enable the bacteria to bind to eukaryotic cells.S. Typhimurium type 1 fimbriae are characterized by mannose-sensitive hemagglutination and are assembled via the chaperone/usher pathway.S. Typhimurium type 1 fimbrial proteins are encoded by thefimgene cluster (fimAICDHFZYW), withfimAICDHFexpressed as a single transcriptional unit. The structural components of the fimbriae are FimA (major subunit), FimI, FimH (adhesin), and FimF (adaptor). In order to determine which components are required for fimbrial formation inS. Typhimurium, mutations infimA,fimI,fimH, andfimFwere constructed and examined for their ability to produce surface-assembled fimbriae.S. Typhimurium SL1344ΔfimA, -ΔfimH, and -ΔfimFmutants were unable to assemble fimbriae, indicating that these genes are necessary for fimbrial production inS. Typhimurium. However, SL1344ΔfimIwas able to assemble fimbriae. InEscherichia colitype 1 and Pap fimbriae, at least two adaptors are expressed in addition to the adhesins. However,E. colitype 1 and Pap fimbriae have been reported to be able to assemble fimbriae in the absence of these proteins. These results suggest differences between theS. Typhimurium type 1 fimbrial system and theE. colitype 1 and Pap fimbrial systems.

Mutations and copy number variants of the CUB and Sushi multiple domains 2 ( CSMD2 ) gene are associated with neuropsychiatric disease. CSMD2 encodes a single-pass transmembrane protein with a large extracellular domain comprising repeats of CUB and Sushi domains. High expression of CSMD2 in the developing and mature brain suggests possible roles in neuron development or function, but the cellular functions of CSMD2 are not known. In this study, we show that mouse Csmd2 is expressed in excitatory and inhibitory neurons in the forebrain. Csmd2 protein exhibits a somatodendritic localization in the neocortex and hippocampus, with smaller puncta localizing to the neuropil. Using immunohistochemical and biochemical methods, we demonstrate that Csmd2 localizes to dendritic spines and is enriched in the postsynaptic density (PSD). Accordingly, we show that the cytoplasmic tail domain of Csmd2 interacts with synaptic scaffolding proteins of the membrane-associated guanylate kinase (MAGUK) family. The association between Csmd2 and MAGUK member PSD-95 is dependent on a PDZ-binding domain on the Csmd2 tail, which is also required for synaptic targeting of Csmd2. Finally, we show that knock-down of Csmd2 expression in hippocampal neuron cultures results in reduced complexity of dendritic arbors and deficits in dendritic spine density. Knock-down of Csmd2 in immature developing neurons results in reduced filopodia density, whereas Csmd2 knock-down in mature neurons causes significant reductions in dendritic spine density and dendrite complexity. Together, these results point toward a function for Csmd2 in development and maintenance of dendrites and synapses, which may account for its association with certain psychiatric disorders.

Type 1 fimbriae produced by serovars of Salmonella are characterized by their ability to agglutinate guinea pig erythrocytes in the absence of D-mannose but not in its presence. The FimH protein is the adhesin that mediates this reaction; it is distinct from the major fimbrial protei.n (FimA) that composes the fimbrial shaft. Avian-adapted serovars of Salmonella produce nonhaemagglutinating fimbriae that have been reported to mediate adherence to avian cells. A single amino acid substitution is present in the FimH adhesin of these strains compared to that of a Typhimurium isolate. Also, previous studies have shown that single nucleotide polymorphisms in two strains of the Typhimurium fimH alter the binding specificity. We therefore investigated the allelic variation of fimH from a range of serotypes (both host-adapted and non-host-adapted) and isolates of Salmonella. Most FimH adhesins mediated the mannose-sensitive haemagglutination of guinea pig erythrocytes, but many did not facilitate adherence to HEp-2 cells. A small number of isolates also produced fimbriae but did not mediate adherence to either cell type. Transformants possessing cloned fimH genes exhibited a number of different substitutions within the predicted amino acid sequence of the FimH polypeptide. No identical FimH amino sequence was found between strains that adhere to erythrocytes and/or HEp-2 cells and those produced by non-adherent strains. FimH-mediated adherence to HEp-2 cells was invariably associated with the ability to form biofilms on mannosylated bovine serum albumin.

During neocortical development, neurons are produced by a diverse pool of neural progenitors. A subset of progenitors express the Cux2 gene and are fate-restricted to produce certain neuronal subtypes, but the upstream pathways that specify these progenitor fates remain unknown. To uncover the transcriptional networks that regulate Cux2 expression in the forebrain, we characterized a conserved Cux2 enhancer that we find recapitulates Cux2 expression specifically in the cortical hem. Using a bioinformatic approach, we found several potential transcription factor (TF) binding sites for cortical hem-patterning TFs. We found that the homeobox transcription factor, Lmx1a, can activate the Cux2 enhancer in vitro. Furthermore, we show that multiple Lmx1a binding sites required for enhancer activity in the cortical hem in vivo. Mis-expression of Lmx1a in neocortical progenitors caused an increase in Cux2+-lineage cells. Finally, we compared several conserved human enhancers with cortical hem-restricted activity and found that recurrent Lmx1a binding sites are a top shared feature. Uncovering the network of TFs involved in regulating Cux2 expression will increase our understanding of the mechanisms pivotal in establishing Cux2-lineage fates in the developing forebrain.Summary StatementAnalysis of a cortical hem-specific Cux2 enhancer reveals role for Lmx1a as a critical upstream regulator of Cux2 expression patterns in neural progenitors during early forebrain development.