Failure of remyelination is largely responsible for sustained neurologic symptoms in multiple sclerosis (MS). MS lesions contain hyaluronan deposits that inhibit oligodendrocyte precursor cell (OPC) maturation. However, the mechanism behind this inhibition is unclear. We report here that Toll-like receptor 2 (TLR2) is expressed by oligodendrocytes and is up-regulated in MS lesions. Pathogenderived TLR2 agonists, but not agonists for other TLRs, inhibit OPC maturation in vitro. Hyaluronan-mediated inhibition of OPC maturation requires TLR2 and MyD88, a TLR2 adaptor molecule. Ablated expression of TLR2 also enhances remyelination in a lysolecithin animal model. Hyaluronidases expressed by OPCs degrade hyaluronan to hyaluronan oligomers, a requirement for hyaluronan/TLR2 signaling. MS lesions contain both TLR2 + oligodendrocytes and lowmolecular-weight hyaluronan, consistent with their importance to remyelination in MS. We thus have defined a mechanism controlling remyelination failure in MS where hyaluronan is degraded by hyaluronidases into hyaluronan oligomers that block OPC maturation and remyelination through TLR2-MyD88 signaling.hyaluronidase | MyD88 | innate immunity I n multiple sclerosis (MS), destruction of CNS myelin accounts for a majority of neurologic symptoms. MS patients typically exhibit a relapsing/remitting course in which relapses result from inflammation and demyelination and remissions result from resolution of inflammation and remyelination. Secondary progressive MS, which typically begins in patients after a decade of relapsing/ remitting MS, exhibits irreversible neurologic disability.Most chronic MS lesions show little if any remyelination. The failure of remyelination in MS theoretically could be the consequence of a deficiency in the number of oligodendrocyte progenitor cells (OPCs), absence of a promyelination signal, or the presence of inhibitory influences on OPCs. It therefore is of interest that the histopathology of the MS lesion has revealed the presence of OPCs and premyelinating oligodendrocytes in chronic MS lesions. The premyelinating oligodendrocytes extend processes that contact but fail to myelinate axons (1-4), thus suggesting failure of remyelination is caused by the loss of promyelination signals or the presence of inhibitory signals.Recently, the glycosaminoglycan hyaluronan was identified within MS lesions and found to inhibit OPC maturation and remyelination in an MS animal model (5). The mechanism underlying this inhibition is unknown. Because hyaluronan can function as an endogenous mammalian ligand for Toll-like receptors (TLRs) 2 and 4 in innate immune cell activation (6-9), we reasoned that TLR stimulation also may be required for hyaluronan-mediated blocking of OPC maturation.Although TLRs and the Drosophila ortholog Toll have welldescribed functions in innate immune cells (6-12), TLRs also have potent functions outside the immune system. Toll and TLR have diverse roles in axonal pathfinding, dorsoventral patterning, and cell-fate determination (13). In particul...

Toll receptors in Drosophila melanogaster function in morphogenesis and host defense. Mammalian orthologues of Toll, the Toll-like receptors (TLRs), have been studied extensively for their essential functions in controlling innate and adaptive immune responses. We report that TLR8 is dynamically expressed during mouse brain development and localizes to neurons and axons. Agonist stimulation of TLR8 in cultured cortical neurons causes inhibition of neurite outgrowth and induces apoptosis in a dissociable manner. Our evidence indicates that such TLR8-mediated neuronal responses do not involve the canonical TLR–NF-κB signaling pathway. These findings reveal novel functions for TLR8 in the mammalian nervous system that are distinct from the classical role of TLRs in immunity.

Toll is a cell surface receptor with well described roles in the developmental patterning of invertebrates and innate immunity in adult Drosophila. Mammalian toll-like receptors represent a family of Toll orthologs that function in innate immunity by recognizing molecular motifs unique to pathogens or injured tissue. One member in this family of pattern recognition receptors, toll-like receptor 3 (TLR3), recognizes viral double-stranded RNA and host mRNA. We examined the expression and function of TLRs in the nervous system and found that TLR3 is expressed in the mouse central and peripheral nervous systems and is concentrated in the growth cones of neurons. Activation of TLR3 by the synthetic ligand polyinosine:polycytidylic acid (poly I:C) or by mRNA rapidly causes growth cone collapse and irreversibly inhibits neurite extension independent of nuclear factor B. Mice lacking functional TLR3 were resistant to the neurodegenerative effects of poly I:C. Neonatal mice injected with poly I:C were found to have fewer axons exiting dorsal root ganglia and displayed related sensorimotor deficits. No effect of poly I:C was observed in mice lacking functional TLR3. Together, these findings provide evidence that an innate immune pattern recognition receptor functions autonomously in neurons to regulate axonal growth and advances a novel hypothesis that this class of receptors may contribute to injury and limited CNS regeneration.

Clostridium perfringens ε-toxin (ETX) is a potent pore-forming toxin responsible for a central nervous system (CNS) disease in ruminant animals with characteristics of blood-brain barrier (BBB) dysfunction and white matter injury. ETX has been proposed as a potential causative agent for Multiple Sclerosis (MS), a human disease that begins with BBB breakdown and injury to myelin forming cells of the CNS. The receptor for ETX is unknown. Here we show that both binding of ETX to mammalian cells and cytotoxicity requires the tetraspan proteolipid Myelin and Lymphocyte protein (MAL). While native Chinese Hamster Ovary (CHO) cells are resistant to ETX, exogenous expression of MAL in CHO cells confers both ETX binding and susceptibility to ETX-mediated cell death. Cells expressing rat MAL are ~100 times more sensitive to ETX than cells expressing similar levels of human MAL. Insertion of the FLAG sequence into the second extracellular loop of MAL abolishes ETX binding and cytotoxicity. ETX is known to bind specifically and with high affinity to intestinal epithelium, renal tubules, brain endothelial cells and myelin. We identify specific binding of ETX to these structures and additionally show binding to retinal microvasculature and the squamous epithelial cells of the sclera in wild-type mice. In contrast, there is a complete absence of ETX binding to tissues from MAL knockout (MAL-/-) mice. Furthermore, MAL-/- mice exhibit complete resistance to ETX at doses in excess of 1000 times the symptomatic dose for wild-type mice. We conclude that MAL is required for both ETX binding and cytotoxicity.

The plant hormone abscisic acid (ABA) play essential roles in numerous physiological processes such as seed dormancy, seed germination, seeding growth and responses to biotic and abiotic stresses. Such biological processes are tightly controlled by a complicated regulatory network including ABA homoeostasis, signal transduction as well as cross-talking among other signaling pathways. It is known that ABA homoeostasis modulated by its production, inactivation, and transport pathways is considered to be of great importance for plant development and stress responses. Most of the enzymes and transporters involved in ABA homoeostasis have been largely characterized and they all work synergistically to maintain ABA level in plants. Increasing evidence have suggested that transcriptional regulation of the genes involved in either ABA production or ABA inactivation plays vital roles in ABA homoeostasis. In addition to transcription factors, such progress is also regulated by microRNAs and newly characterized root to shoot mobile peptide-receptor like kinase (RLKs) mediated long-distance signal transduction. Thus, ABA contents are always kept in a dynamic balance. In this review, we survey recent research on ABA production, inactivation and transport pathways, and summarize some latest findings about the mechanisms that regulate ABA homoeostasis.

SUMMARY The molecular pathways controlling cerebellar Purkinje cell dendrite formation and maturation are poorly understood. The Purkinje cell degeneration (pcd) mutant mouse is characterized by mutations in Nna1a gene discovered in an axonal regenerative context, but whose actual function in development and disease is unknown. We found abnormal development of Purkinje cell dendrites in postnatal pcdSid mice and linked this deficit to a deletion mutation in exon 7 of Nna1. With single cell gene profiling and virus-based gene transfer, we analyzed a molecular pathway downstream to Nna1 underlying abnormal Purkinje cell dendritogenesis in pcdSid mice. We discovered that mutant Nna1 dramatically increases intranuclear localization of lysyl oxidase propeptide, which interferes with NF-кB RelA signaling and microtubule-associated protein regulation of microtubule stability, leading to underdevelopment of Purkinje cell dendrites. These findings provide insight into Nna1’s role in neuronal development and why its absence renders Purkinje cells more vulnerable.

Clostridium perfringens epsilon toxin (ε-toxin) is responsible for a devastating multifocal central nervous system (CNS) white matter disease in ruminant animals. The mechanism by which ε-toxin causes white matter damage is poorly understood. In this study, we sought to determine the molecular and cellular mechanisms by which ε-toxin causes pathological changes to white matter. In primary CNS cultures, ε-toxin binds to and kills oligodendrocytes but not astrocytes, microglia, or neurons. In cerebellar organotypic culture, ε-toxin induces demyelination, which occurs in a time- and dose-dependent manner, while preserving neurons, astrocytes, and microglia. ε-Toxin specificity for oligodendrocytes was confirmed using enriched glial culture. Sensitivity to ε-toxin is developmentally regulated, as only mature oligodendrocytes are susceptible to ε-toxin; oligodendrocyte progenitor cells are not. ε-Toxin sensitivity is also dependent on oligodendrocyte expression of the proteolipid myelin and lymphocyte protein (MAL), as MAL-deficient oligodendrocytes are insensitive to ε-toxin. In addition, ε-toxin binding to white matter follows the spatial and temporal pattern of MAL expression. A neutralizing antibody against ε-toxin inhibits oligodendrocyte death and demyelination. This study provides several novel insights into the action of ε-toxin in the CNS. (i) ε-Toxin causes selective oligodendrocyte death while preserving all other neural elements. (ii) ε-Toxin-mediated oligodendrocyte death is a cell autonomous effect. (iii) The effects of ε-toxin on the oligodendrocyte lineage are restricted to mature oligodendrocytes. (iv) Expression of the developmentally regulated proteolipid MAL is required for the cytotoxic effects. (v) The cytotoxic effects of ε-toxin can be abrogated by an ε-toxin neutralizing antibody.

AbbReViATionSIκBα nuclear factor of κ light polypeptide gene enhancer in B-cells inhibitor α IRAK interleukin-1 receptor-associated kinase LPS lipopolysaccharide MyD88 myeloid differentiation factor 88 NFκB nuclear factor-κB PAMPs pathogen-associated molecular patterns TLR Toll-like receptor AcKnowLedgeMenTSWe thank colleagues I. Chiu, J. Lu, and J. Sloane for critical reading of the manuscript. This work was supported by grants from the US National Institutes of Health and the National Multiple Sclerosis Society. AbSTRAcTToll-like receptors (TLRs) play essential roles in generating innate immune responses, and are evolutionarily conserved across species. In mammals, TLRs specifically recognize the conserved microbial structural motifs referred to as pathogen-associated molecular patterns (PAMPs). Ligand recognition by TLRs activates signaling cascades that culminate in proinflammatory cytokine production and eventual elimination of invading pathogens. Although TLRs in mammals are expressed predominantly in the immune system, certain TLRs with poorly characterized function are also found in neurons. We recently profiled TLR8 expression during mouse brain development and established its localization in neurons and axons. We uncovered a novel role for TLR8 as a suppressor of neurite outgrowth as well as an inducer of neuronal apoptosis, and found that TLR8 functions in neurons through an NFκB-independent mechanism. These findings add a new layer of complexity for TLR signaling, and expand the realm of mammalian TLR function to the central nervous system (CNS) beyond the originally discovered immune context. Herein, we complement our earlier report with additional data, discuss their biological and mechanistic implications in CNS developmental and pathological processes, and thus further our perspective on TLR signaling and potential physiological roles in mammals.