[Show abstract][Hide abstract] ABSTRACT: The oligodendrocyte lineage genes Olig1 and Olig2 encode related bHLH proteins that are co-expressed in neural progenitors. Targeted disruption of these two genes sheds light on the ontogeny of oligodendroglia and genetic requirements for their development from multipotent CNS progenitors. Olig2 is required for oligodendrocyte and motor neuron specification in the spinal cord. Olig1 has roles in development and maturation of oligodendrocytes, evident especially within the brain. Both Olig genes contribute to neural pattern formation. Neither Olig gene is required for astrocytes. These findings, together with fate mapping of Olig-expressing cells, challenge the notion that oligodendrocytes arise from a glial-restricted precursor. Rather, they indicate that oligodendrocytes are derived from Olig-specified progenitors that give rise also to neurons.
Journal of Neurochemistry 06/2008; 81:75-75. · 3.97 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Oligodendrocyte precursor development in the embryonic spinal cord is thought to be regulated by the secreted signal, Sonic hedgehog (Shh). Such precursors can be identified by the expression of Olig genes, encoding basic helix-loop-helix factors, in the spinal cord and brain. However, the signaling pathways that govern oligodendrocyte precursor (OLP) development in the rostral central nervous system are poorly understood. Here, we show that Shh is required for oligodendrocyte development in the mouse forebrain and spinal cord, and that Shh proteins are both necessary and sufficient for OLP production in cortical neuroepithelial cultures. Moreover, adenovirus-mediated Olig1 ectopic expression can promote OLP formation independent of Shh activity. Our results demonstrate essential functions for Shh during early phases of oligodendrocyte development in the mammalian central nervous system. They further suggest that a key role of Shh signaling is activation of Olig genes.
Molecular and Cellular Neuroscience 11/2001; 18(4):434-41. · 3.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The most common primary tumors of the human brain are thought to be of glial cell origin. However, glial cell neoplasms cannot be fully classified by cellular morphology or with conventional markers for astrocytes, oligodendrocytes, or their progenitors. Recent insights into central nervous system tumorigenesis suggest that novel molecular markers might be found among factors that have roles in glial development. Oligodendrocyte lineage genes (Olig1/2) encode basic helix-loop-helix transcription factors. In the rodent central nervous system, they are expressed exclusively in oligodendrocytes and oligodendrocyte progenitors, and Olig1 can promote formation of an chondroitin sulfate proteoglycon-positive glial progenitor. Here we show that human OLIG genes are expressed strongly in oligodendroglioma, contrasting absent or low expression in astrocytoma. Our data provide evidence that neoplastic cells of oligodendroglioma resemble oligodendrocytes or their progenitor cells and may derive from cells of this lineage. They further suggest the diagnostic potential of OLIG markers to augment identification of oligodendroglial tumors.
Proceedings of the National Academy of Sciences 10/2001; 98(19):10851-6. · 9.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Organizing signals such as Sonic hedgehog are thought to specify neuronal subtype identity by regulating the expression of homeodomain proteins in progenitors of the embryonic neural tube. One of these, Nkx2.2, is necessary and sufficient for the development of V3 interneurons.
We report that Olig genes, encoding basic helix-loop-helix (bHLH) proteins, are expressed in a subset of Nkx2.2 progenitors before the establishment of interneurons and oligodendroglial precursors. Gain-of-function analysis in transgenic mouse embryos indicates that Olig genes specifically inhibit the establishment of Sim1-expressing V3 interneurons. Moreover, coexpression of Olig2 with Nkx2.2 in the chick neural tube generated cells expressing Sox10, a marker of oligodendroglial precursors. Colocalization of Olig and Nkx2.2 proteins at the dorsal extent of the Nkx2.2 expression domain is consistent with regulatory interactions that define the potential of progenitor cells in the border region.
Interactions between homeodomain and Olig bHLH proteins evidently regulate neural cell fate acquisition and diversification in the ventral neural tube. In particular, interactions between Olig and Nkx2.2 proteins inhibit V3 interneuron development and promote the formation of alternate cell types, including those expressing Sox10.
Current Biology 10/2001; 11(18):1413-20. · 9.49 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: There are clear parallels between oligodendrocyte development in the spinal cord and forebrain. However, there is new evidence that in both of these regions oligodendrocyte lineage development may be more complex than we earlier thought. This stems from the recent identification of three new transcription factor genes, Olig1, Olig2 and Sox10, that are expressed from the early stages of oligodendrocyte lineage development. In this article, we highlight the common themes underlying specification and early development of oligodendrocytes in the spinal cord and telencephalon. Then, we discuss recent studies of Sox10 and the Olig genes and their implications for oligodendrocyte specification. We conclude that although the mechanisms of oligodendrogenesis appear to be fundamentally similar at different rostro-caudal levels of the neuraxis, there are still many unanswered questions about the details of oligodendrocyte specification.
International Journal of Developmental Neuroscience 08/2001; 19(4):379-85. · 2.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In the caudal neural tube, oligodendrocyte progenitors (OLPs) originate in the ventral neuroepithelium under the influence of Sonic hedgehog (SHH), then migrate throughout the spinal cord and brainstem before differentiating into myelin-forming cells. We present evidence that oligodendrogenesis in the anterior neural tube follows a similar pattern. We show that OLPs in the embryonic mouse forebrain express platelet-derived growth factor alpha-receptors (PDGFRA), as they do in more caudal regions. They first appear within a region of anterior hypothalamic neuroepithelium that co-expresses mRNA encoding SHH, its receptor PTC1 (PTCH) and the transcription factors OLIG1, OLIG2 and SOX10. Pdgfra-positive progenitors later spread through the forebrain into areas where Shh is not expressed, including the cerebral cortex. Cyclopamine inhibited OLP development in cultures of mouse basal forebrain, suggesting that hedgehog (HH) signalling is obligatory for oligodendrogenesis in the ventral telencephalon. Moreover, Pdgfra-positive progenitors did not appear on schedule in the ventral forebrains of Nkx2.1 null mice, which lack the telencephalic domain of Shh expression. However, OLPs did develop in cultures of Nkx2.1(-/-) basal forebrain and this was blocked by cyclopamine. OLPs also developed in neocortical cultures, even though Shh transcripts could not be detected in the embryonic cortex. Here, too, the appearance of OLPs was suppressed by cyclopamine. In keeping with these findings, we detected mRNA encoding SHH and Indian hedgehog (IHH) in both Nkx2.1(-/-) basal forebrain cultures and neocortical cultures. Overall, the data are consistent with the idea that OLPs in the telencephalon, possibly even some of those in the cortex, develop under the influence of SHH in the ventral forebrain.
Development 08/2001; 128(13):2545-54. · 6.21 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In most mammalian cells, the cAMP-dependent protein kinase A pathway promotes growth arrest and cell differentiation. However in Schwann cells, the reverse is true. Elevated levels of cAMP function as the cofactor to a broad range of mitogenic cues in culture and in animals. Previous studies have suggested that cAMP acts at an early point in the Schwann cell mitogenic response, perhaps by stimulating the expression of growth factor receptors. We show here that cAMP acts downstream rather than upstream of growth factor receptor expression. The essential function(s) of cAMP is exerted as Schwann cells progress through the G(1) phase of the cell cycle. Ectopic expression studies using an inducible retroviral vector show that the G(1) phase requirement for cAMP can be alleviated by a single protein, cyclin D1. We show, in addition, that at least one function of the Nf1 tumor suppressor is to antagonize the accumulation of cAMP and the expression of cyclin D1 in Schwann cells. Thus a G(1) phase-specific protein, cyclin D1, accounts for two salient features of Schwann cell growth control: the promitotic response to cAMP and the antimitotic response to the Nf1 tumor suppressor.
Journal of Neuroscience 03/2001; 21(4):1110-6. · 6.91 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Glioblastoma multiforme is the most common primary human brain tumor, and it is, for all practical purposes, incurable in adult patients. The high mortality rates reflect the fact that glioblastomas are resistant to adjuvant therapies (radiation and chemicals), the mode of action of which is cytotoxic. We show here that an p.o.-active small molecule kinase inhibitor of the 2-phenylaminopyrimidine class may have therapeutic potential for glioblastomas. STI571 inhibits the growth of U343 and U87 human glioblastoma cells that have been injected into the brains of nude mice, but it does not inhibit intracranial growth of ras-transformed cells. Studies on a broad panel of genetically validated human and animal cell lines show that STI571 acts by disruption of the ligand:receptor autocrine loops for platelet-derived growth factor that are a pervasive feature of malignant astrocytoma. The cellular response of glioblastoma cells to STI571 does not appear to involve an apoptotic mechanism.
Cancer Research 10/2000; 60(18):5143-50. · 8.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Sciatic nerve axons in cyclin D1 knockout mice develop normally, become properly ensheathed by Schwann cells, and appear to function normally. However, in the Wallerian degeneration model of nerve injury, the mitotic response of Schwann cells is completely inhibited. The mitotic block is Schwann cell autonomous and developmentally regulated. Rescue analysis (by "knockin" of cyclin E) indicates that D1 protein, rather than regulatory elements of the D1 gene, provides the essential Schwann cell function. Genetic inhibition of the Schwann cell cycle shows that neuronal responses to nerve injury are surprisingly independent of Schwann cell mitotic responses. Even axonal regrowth into the distal zone of a nerve crush injury is not markedly impaired in cyclin D1-/- mice.
[Show abstract][Hide abstract] ABSTRACT: During development, basic helix-loop-helix (bHLH) proteins regulate formation of neurons from multipotent progenitor cells. However, bHLH factors linked to gliogenesis have not been described. We have isolated a pair of oligodendrocyte lineage genes (Olg-1 and Olg-2) that encode bHLH proteins and are tightly associated with development of oligodendrocytes in the vertebrate central nervous system (CNS). Ectopic expression of Olg-1 in rat cortical progenitor cell cultures promotes formation of oligodendrocyte precursors. In developing mouse embryos, Olg gene expression overlaps but precedes the earliest known markers of the oligodendrocyte lineage. Olg genes are expressed at the telencephalon-diencephalon border and adjacent to the floor plate, a source of the secreted signaling molecule Sonic hedgehog (Shh). Gain- and loss-of-function analyses in transgenic mice demonstrate that Shh is both necessary and sufficient for Olg gene expression in vivo.
[Show abstract][Hide abstract] ABSTRACT: Neurons and glia of the cerebral cortex are thought to arise from a common, multipotent progenitor cell that is instructed toward alternate fates by extracellular cues. How do these cells behave when confronted with conflicting cues? We show here that nestin-positive neuroepithelial (NE) cells from embryonic day 14 rat cortex coexpress surface receptor proteins for ciliary neurotrophic factor (CNTF) and platelet-derived growth factor (PDGF). Both sets of these receptor proteins are functional in NE cells, as shown by ligand-dependent activation of downstream signal-generating proteins. Transient (30') exposure to CNTF instructs NE cells toward an astrocyte fate. Brief exposure to PDGF initiates neuronal differentiation. However, when challenged with conflicting cues, PDGF is dominant to CNTF. Moreover, CNTF-treated NE cells can be "redirected" by a subsequent exposure to PDGF to form neurons instead of astrocytes, whereas the converse is not true. The asymmetric relationship between CNTF and PDGF indicates that these two growth factors act on a common progenitor cell that has, at a minimum, two fates available to it rather than separate populations of precommitted neuroblasts and astroblasts. This bipotent progenitor cell processes conflicting cues for neurons and glia in a hierarchical manner.
Journal of Neuroscience 01/2000; 19(23):10383-9. · 6.91 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Platelet-derived growth factor (PDGF) stimulates transcription of an immediate-early gene set in Balb/c 3T3 cells. One cohort of these genes, typified by c-fos, is induced within minutes following activation of PDGF receptors. A second cohort responds to PDGF only after a significant time delay, although induction is still a primary response to receptor activation as shown by "superinduction" in the presence of the protein synthesis inhibitor cycloheximide. PDGF-receptor activated signaling pathways for the "slow" immediate-early genes are poorly resolved. Using gain-of-function mutations together with small molecule inhibitors of kinase activity, we show that activation of PI 3-kinase is both necessary and sufficient for the induction of the prototype slow immediate-early gene, monocyte chemoattractant-1 (MCP-1). Following activation of PDGF receptors, MCP-1 mRNA does not begin to accumulate for at least 90 min. However, only a brief (10 min) interval of PI 3-kinase activity is required to trigger this delayed response. The serine/threonine protein kinase, Akt/PKB, likely functions as a downstream affector of PI 3-kinase for this induction.
Journal of Biological Chemistry 11/1999; 274(43):31062-7. · 4.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The human nerve growth factor receptor (TrkA) contains four potential N-glycosylation sites that are highly conserved within the Trk family of neurotrophin receptors, and nine additional sites that are less well conserved. Using a microscale deglycosylation assay, we show here that both conserved and variable N-glycosylation sites are used during maturation of TrkA. Glycosylation at these sites serves two distinct functions. First, glycosylation is necessary to prevent ligand-independent activation of TrkA. Unglycosylated TrkA core protein is phosphorylated even in the absence of ligand stimulation and displays constitutive kinase activity as well as constitutive interaction with the signaling molecules Shc and PLC-gamma. Second, glycosylation is required to localize TrkA to the cell surface, where it can trigger the Ras/Raf/MAP kinase cascade. Using confocal microscopy, we show that unglycosylated active Trk receptors are trapped intracellularly. Furthermore, the unglycosylated active TrkA receptors are unable to activate kinases in the Ras-MAP kinase pathway, MEK and Erk. Consistent with these biochemical observations, unglycosylated TrkA core protein does not promote neuronal differentiation in Trk PC12 cells even at high levels of constitutive catalytic activity.
Journal of Neurobiology 06/1999; 39(2):323-36. · 3.05 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: PTEN is a novel tumour suppressor gene that encodes a dual-specificity phosphatase with homology to adhesion molecules tensin and auxillin. It recently has been suggested that PTEN dephosphorylates phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3, 4,5)P3], which mediates growth factor-induced activation of intracellular signalling, in particular through the serine-threonine kinase Akt, a known cell survival-promoting factor. PTEN has been mapped to 10q23.3, a region disrupted in several human tumours including haematological malignancies. We have analysed PTEN in a series of primary acute leukaemias and non-Hodgkin's lymphomas (NHLs) as well as in cell lines. We have also examined whether a correlation could be found between PTEN and Akt levels in these samples. We show here that the majority of cell lines studied carries PTEN abnormalities. At the structural level, we found mutations and hemizygous deletions in 40% of these cell lines, while a smaller number of primary haematological malignancies, in particular NHLs, carries PTEN mutations. Moreover, one-third of the cell lines had low PTEN transcript levels, and 60% of these samples had low or absent PTEN protein, which could not be attributed to gene silencing by hypermethylation. In addition, we found that PTEN and phosphorylated Akt levels are inversely correlated in the large majority of the examined samples. These findings suggest that PTEN plays a role in the pathogenesis of haematological malignancies and that it might be inactivated through a wider range of mechanisms than initially considered. The finding that PTEN levels inversely correlate with phosphorylated Akt supports the hypothesis that PTEN regulates PtdIns(3,4,5)P3and suggests a role for PTEN in apoptosis.
Human Molecular Genetics 03/1999; 8(2):185-93. · 7.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Elevated expression of the neurotrophin-3 (NT-3) receptor TrkC by childhood medulloblastomas is associated with favorable clinical outcome. Here, we provide evidence that TrkC is more than simply a passive marker of prognosis. We demonstrate that: (a) medulloblastomas undergo apoptosis in vitro when grown in the presence of NT-3; (b) overexpression of TrkC inhibits the growth of intracerebral xenografts of a medulloblastoma cell line in nude mice; and (c) trkC expression by individual tumor cells is highly correlated with apoptosis within primary medulloblastoma biopsy specimens. TrkC-mediated NT-3 signaling promotes apoptosis by activating multiple parallel signaling pathways and by inducing immediate-early gene expression of both c-jun and c-fos. Considered collectively, these results support the conclusion that the biological actions of TrkC activation affect medulloblastoma outcome by inhibiting tumor growth through the promotion of apoptosis.
Cancer Research 03/1999; 59(3):711-9. · 8.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We used anti-phosphopeptide-immunodetecting antibodies as immunohistochemical reagents to define the location and activity state of p185(erbB2) during Wallerian degeneration. Nerve damage induces a phosphorylation event at Y1248, a site that couples p185(erbB2) to the Ras-Raf-MAP kinase signal transduction pathway. Phosphorylation of p185(erbB2) occurs within Schwann cells and coincides in time and space with Schwann cell mitotic activity, as measured by bromodeoxyuridine uptake. These visual images of receptor autophosphorylation link activation of p185(erbB2) to the Schwann cell proliferation that accompanies nerve regeneration.
Journal of Neuroscience 12/1997; 17(21):8293-9. · 6.91 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The beta receptor subunit of platelet-derived growth factor (PDGF) and its corresponding ligand (PDGF-BB) are coordinately expressed in fresh surgical isolates of human meningioma. These observations imply that PDGF autocrine loops are engaged in human meningioma and suggest that activated PDGF-beta receptors might contribute to the pathology of this common brain neoplasm. The study of PDGF autocrine loops and human meningioma has been slowed by the scarcity of meningioma cell culture model systems. Furthermore, in meningioma tumor tissue, the activation state of PDGF receptors is difficult to assess with conventional reagents, because the tumor is intermixed with normal stroma. In fact, there is no evidence that PDGF receptors within the tumor are activated by ligand. We used a synthetic tyrosine phosphopeptide to raise an antibody that reports the phosphorylation state of tyrosine 751 in the human PDGF-beta receptor. Phosphorylated tyrosine 751 is a recognition site for phosphatidylinositol 3'-kinase, a cytoplasmic effector of PDGF-induced mitogenesis, chemotaxis, and membrane ruffling. Immunoblotting and immunostaining analyses with this antibody show that the PDGF-beta receptor is constitutively phosphorylated at tyrosine 751 within multiple fresh surgical isolates of human meningioma. These findings are consistent with a role for activated PDGF receptors in the proliferation of human meningiomas.
Cancer Research 10/1997; 57(18):4141-7. · 8.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Synthetic tyrosine phosphopeptides were used to generate antibodies that recognize phosphotyrosine in the context of a defined sequence of flanking amino acids. Using phosphopeptide immunogens derived from regulatory or signal-generating motifs, "phosphorylation-directed antibodies" can be targeted to specific growth factor receptors or signal-generating proteins. In his paper, we show how phosphorylation-directed antibodies can be used in a colorimetric, high-throughput screen for drugs that modulate the function of specific growth factor receptors or signal-generating proteins.
[Show abstract][Hide abstract] ABSTRACT: During development target-derived neurotrophins promote the survival of neurons. However, mature neurons no longer depend on the target for survival. Do target-derived neurotrophins retain retrograde signaling functions in mature neurons, and, if so, how are they executed? We addressed this question by using a phosphotyrosine-directed antibody to locate activated Trk receptors in adult rat sciatic nerve. We show that catalytically active Trk receptors are located within the axon of adult rat sciatic nerve and that they are distributed throughout the length of the axons. These catalytically active receptors are phosphorylated on tyrosine at a position that couples them to the signal-generating proteins Ras and PI3 kinase. Neurotrophin applied at sciatic nerve terminals increases both catalytic activity and phosphorylation state of Trk receptors at distant points within the axons. Trk activation initiated at the nerve terminals propagates through the axon toward the nerve cell body at an initial rate that exceeds that of conventional vesicular transport. However, our data suggest that this rapid signal is nevertheless vesicle-associated. Thus, in mature nerves, activated Trk receptors function as rapid retrograde signal carriers to execute remote responses to target-derived neurotrophins.
Journal of Neuroscience 10/1997; 17(18):7007-16. · 6.91 Impact Factor