Low-threshold mechanoreceptor neurons (LTMs) of the dorsal root ganglia (DRG) are essential for touch sensation. They form highly specialized terminations in the skin and display stereotyped ...projections in the spinal cord. Functionally defined LTMs depend on neurotrophin signaling for their postnatal survival and functioning, but how these neurons arise during development is unknown. Here, we show that specific types of LTMs can be identified shortly after DRG genesis by unique expression of the MafA transcription factor, the Ret receptor and coreceptor GFRα2, and find that their specification is Ngn2 dependent. In mice lacking Ret, these LTMs display early differentiation defects, as revealed by reduced MafA expression, and at later stages their central and peripheral projections are compromised. Moreover, in MafA mutants, a discrete subset of LTMs display altered expression of neurotrophic factor receptors. Our results provide evidence that genetic interactions involving Ret and MafA progressively promote the differentiation and diversification of LTMs.
We report the identification of an additional member of the glial cell line-derived neurotrophic factor (GDNF) family receptor, termed GFRα3, that is homologous to the previously identified GDNF and ...neurturin ligand binding receptors GFRα1 and GFRα2. GFRα3 is 32% and 37% identical to GFRα1 and GFRα2, respectively. RNase protection assays show that whereas gfrα1 and gfrα2 are abundant in both developing and adult brain, gfrα3 is exclusively expressed during development. All receptors are widely present in both the developing and adult peripheral nervous system and in peripheral organs. For instance, in situ hybridization shows that the developing liver, stomach, intestine, kidney, and sympathetic chain, which all contain ret-expressing cells, transcribe unique complementary and overlapping patterns of most or all of the GDNF family receptors and ligands. In sensory neurons of the trigeminal ganglion gfrα2 and gfrα3 are expressed in different subpopulations of neurons, whereas gfrα1 is coexpressed in some gfrα2 and gfrα3-positive neurons. We find that the gfrα1 population of trigeminal neurons is absent in GDNF null mutant mice, suggesting that GDNF signals in vivo by interacting with GFRα1. Thus, our results show that there are at least three members in the GDNF family of ligand binding receptors and that these receptors may be crucial in conferring ligand specificity in vivo . The unique complementary and overlapping expression of gfrα3 implies distinct functions in the developing and adult mouse from that of GFRα1 and GFRα2.
Glial cell line-derived neurotrophic factor (GDNF) promotes the survival of postnatal-but not embryonic-mouse dorsal root ganglion cells in vitro, despite the fact that its receptors are expressed at ...both ages. To understand this difference, we have performed an oligonucleotide microarray experiment. We found that several hundred genes were regulated between embryonic and postnatal stages, and that several important classes of genes were differentially regulated by GDNF treatment, including genes related to translation and to phenotypic specification and maturation. Interestingly, a set of genes related to cell adhesion, cytoskeleton and cellular morphology were consistently down-regulated by GDNF, suggesting a previously uncharacterized role for GDNF in repressing neurite growth and/or branching. This nuclear program initiated by GDNF was functionally confirmed in cultures of embryonic wild-type neurons sustained with nerve growth factor and in bax-/-neurons that survive in the absence of trophic support.
Establishment of the neuromuscular synapse requires bidirectional signaling between the nerve and muscle. Although much is known on nerve-released signals onto the muscle, less is known of signals ...important for presynaptic maturation of the nerve terminal. Our results suggest that the Ret tyrosine kinase receptor transmits a signal in motor neuron synapses that contribute to motor neuron survival and synapse maturation at postnatal stages. Ret is localized specifically to the presynaptic membrane with its ligands, GDNF (glial cell line-derived neurotrophic factor)/NTN (neurturin), expressed in skeletal muscle tissue. Lack of Ret conditionally in cranial motor neurons results in a developmental deficit of maturation and specialization of presynaptic neuromuscular terminals. Regeneration of Ret-deficient adult hypoglossal motor neurons is unperturbed, but despite contact with the unaffected postsynaptic specializations, presynaptic axon terminal maturation is severely compromised in the absence of Ret signaling. Thus, Ret transmits a signal in motor nerve terminals that participate in the organization and maturation of presynaptic specializations during development and during regeneration in the adult.
Mantle cell lymphoma (MCL) is a B-cell malignancy characterized by a monoclonal proliferation of lymphocytes with the co-expression of CD5 and CD43, but not of CD23. Typical MCL is associated with ...overexpression of cyclin D1, and blastoid MCL variants are associated with Myc (alias c- myc ) translocations. In this study, we developed a murine model of MCL-like lymphoma by crossing Cdk4 R24C mice with Myc-3′RR transgenic mice. The Cdk4 R24C mouse is a knockin strain that expresses a Cdk4 protein that is resistant to inhibition by p16INK4a as well as other INK4 family members. Ablation of INK4 control on Cdk4 does not affect lymphomagenesis, B-cell maturation, and functions in Cdk4 R24C mice. Additionally, B cells were normal in numbers, cell cycle activity, mitogen responsiveness, and Ig synthesis in response to activation. By contrast, breeding Cdk4 R24C mice with Myc-3′RR transgenic mice prone to develop aggressive Burkitt lymphoma–like lymphoma (CD19+ IgM+ IgD+ cells) leads to the development of clonal blastoid MCL-like lymphoma (CD19+ IgM+ CD5+ CD43+ CD23− cells) in Myc / Cdk4 R24C mice. Western blot analysis revealed high amounts of Cdk4/cyclin D1 complexes as the main hallmark of these lymphomas. These results indicate that although silent in nonmalignant B cells, a defect in the INK4-Cdk4 checkpoint can participate in lymphomagenesis in conjunction with additional alterations of cell cycle control, a situation that might be reminiscent of the development of human blastoid MCL.
Newborn rat brain astrocytes, cultured in a serum-free medium, were exposed for 30 min to two types of reactive oxygen species. Cells were either treated with the xanthine/xanthine oxidase (X/XOD) ...system, which generates both H2O2 and the O2.- radical, or to H2O2 alone. Both treatments induced a dose-dependent accumulation of nerve growth factor (NGF) transcripts, 6 h after the exposure. Maximal effect was obtained with 6 mU/ml XOD, or 10(-4) M H2O2. A rapid expression of protooncogenes of the jun and fos families was also noticed in X/XOD- or H2O2-treated cells. This phenomenon was transient in cells exposed to X/XOD. However, in the case of H2O2-treated cells, the accumulation of c-fos or c-jun mRNAs was still pronounced 6 h after the end of the treatment and the levels of cell-secreted NGF appeared relatively reduced, when compared with those obtained after a shock with the X/XOD system. This raised the possibility that H2O2 at 10(-4) M could depress protein synthesis. Measurements of the incorporation of radiolabeled amino acids into trichloroacetic acid-precipitable material supported this assumption. Level of radioactivity associated with cellular material was dramatically reduced in H2O2-treated cells, when it was compared with control or X/XOD-treated cells. Furthermore, treatment of cells with the protein synthesis inhibitor anisomycin had an effect similar to that of H2O2 because it caused an accumulation of c-fos, c-jun, and NGF transcripts after 6 h of treatment.
Peripheral sensory neurons are derived from two distinct structures, the ectodermal placodes and the neural crest. Here, we establish the forkhead family transcription factor Foxs1 as an early ...sensory neuronal marker. Early embryonic Foxs1 expression was present in all the sensory nervous system regardless of cellular origin, but was not found in other placode and neural crest-derived cell types. Foxs1 expression was turned on in the sensory neuron precursors of the trunk. Consistently, expression of Sox10, that is present in undifferentiated multipotent neural crest cells (NCCs), was mutually exclusive to Foxs1. Acquirement of Foxs1 expression was used to study the emergence of the dorsal root ganglion (DRG) sensory neurons. Migrating pioneering Foxs1 expressing NCCs were found at the anterior dorsal somitic lip at the 18-somite stage. These cells showed limited proliferation and migrated to form a cluster in the ventral aspect of the coalescing ganglion, surrounded by Foxs1
−/Sox10
+ migrating NCCs retaining a high rate of proliferation. Sensory neurogenesis of the Foxs1
−/Sox10
+ precursors occurred within the condensed DRG starting with neurogenin-1 (Ngn1) and Brn3a expression. These data define a sequential emergence of neuronal precursors of the sensory nervous system with different molecular characteristics, starting during migration and continuing well after DRG condensation.
Glial cell line-derived neurotrophic factor (GDNF), neurturin (NTN) and neublastin/artemin (ART) are distant members of the transforming growth factor beta family, and have been shown to elicit ...neurotrophic effects upon several classes of peripheral and central neurons. Limited information from in vitro and expression studies has also substantiated a role for GDNF family ligands in mammalian somatosensory neuron development. Here, we show that although dorsal root ganglion (DRG) sensory neurons express GDNF family receptors embryonically, they do not survive in response to their ligands. The regulation of survival emerges postnatally for all GDNF family ligands. GDNF and NTN support distinct subpopulations that can be separated with respect to their expression of GDNF family receptors, whereas ART supports neurons in populations that are also responsive to GDNF or NTN. Sensory neurons that coexpress GDNF family receptors are medium sized, whereas small-caliber nociceptive cells preferentially express a single receptor. In contrast to brain-derived neurotrophic factor (BDNF)-dependent neurons, embryonic nerve growth factor (NGF)-dependent nociceptive neurons switch dependency to GDNF, NTN and ART postnatally. Neurons that survive in the presence of neurotrophin 3 (NT3) or neurotrophin 4 (NT4), including proprioceptive afferents, Merkel end organs and D-hair afferents, are also supported by GDNF family ligands neonatally, although at postnatal stages they lose their dependency on GDNF and NTN. At late postnatal stages, ART prevents survival elicited by GDNF and NTN. These data provide new insights on the roles of GDNF family ligands in sensory neuron development.
We report the identification of an additional member of the glial cell line-derived neurotrophic factor (GDNF) family receptor, termed GFRα 3, that is homologous to the previously identified GDNF and ...neurturin ligand binding receptors GFRα 1 and GFRα 2. GFRα 3 is 32% and 37% identical to GFRα 1 and GFRα 2, respectively. RNase protection assays show that whereas gfrα 1 and gfrα 2 are abundant in both developing and adult brain, gfrα 3 is exclusively expressed during development. All receptors are widely present in both the developing and adult peripheral nervous system and in peripheral organs. For instance, in situ hybridization shows that the developing liver, stomach, intestine, kidney, and sympathetic chain, which all contain ret-expressing cells, transcribe unique complementary and overlapping patterns of most or all of the GDNF family receptors and ligands. In sensory neurons of the trigeminal ganglion gfrα 2 and gfrα 3 are expressed in different subpopulations of neurons, whereas gfrα 1 is coexpressed in some gfrα 2 and gfrα 3-positive neurons. We find that the gfrα 1 population of trigeminal neurons is absent in GDNF null mutant mice, suggesting that GDNF signals in vivo by interacting with gfrα 1. Thus, our results show that there are at least three members in the GDNF family of ligand binding receptors and that these receptors may be crucial in conferring ligand specificity in vivo. The unique complementary and overlapping expression of gfrα 3 implies distinct functions in the developing and adult mouse from that of GFRα 1 and GFRα 2.
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