C-ret mRNA Research Articles

Discordant expression of c-Ret and glial cell line-derived neurotrophic factor receptor alpha-1 mRNAs in response to motor nerve injury in neonate rats

Adult motoneurons can survive following axotomy, whereas neonate motoneurons result in cell death. Following hypoglossal nerve axotomy in neonate rat, Glial cell line-Derived Neurotrophic Factor (GDNF) receptor alpha-1 (GFRα-1) mRNA expression was dramatically suppressed in the injured motoneurons, while a slight increase of c-Ret mRNA expression was observed. In adult, both GFRα-1 and c-Ret mRNAs increased substantially after axotomy. The present result suggests that the difference of motoneuron fate after axotomy may be partly due to the coordinate or discordant responses of GFRα-1 and c-Ret expression to nerve injury.

GDNF family ligands and receptors are differentially regulated after brain insults in the rat

Expression of mRNAs for glial cell line-derived neurotrophic factor (GDNF), neurturin (NTN) and their receptors was studied in adult rat brain using in situ hybridization after 40 kindling-evoked, rapidly recurring seizures or 10 min of global forebrain ischaemia. Following seizures, GDNF and NTN mRNAs were elevated in dentate granule cells, and c-Ret mRNA in hilar neurons and non-pyramidal cells in CA1 and CA3 regions. GFRalpha-1 mRNA levels showed more widespread increases in the dentate granule cell layer and hilus, CA1 and CA3 pyramidal layers, basolateral amygdala and parietal cortex. The expression of GFRalpha-2 mRNA increased in the piriform cortex and decreased in the CA1 region and basolateral amygdala. Forebrain ischaemia induced elevated expression of GDNF mRNA in dentate granule cells, GFRalpha-1 mRNA in the dentate granule cell layer, hilus and CA3 pyramidal layer, and GFRalpha-2 mRNA in the parietal cortex. The gene expression patterns observed here suggest that GDNF and NTN may act as target-derived factors, but also in an autocrine or paracrine manner. GFRalpha-1 can be coexpressed with GFRalpha-2 and c-Ret mRNAs in the same hippocampal or thalamic neurons, but other neurons contain GFRalpha-1 alone or together with c-Ret mRNA. The gene expression changes for the ligands, and the receptor components are region-, cell- and insult-specific, and occur independently of each other, mainly within 24 h after seizures or ischaemia. This dynamic regulation of GDNF and NTN circuits primarily at the receptor level may be important for the effectiveness of neuroprotective responses but could also trigger plastic changes, e.g. those underlying the development of epileptic syndromes.

Regulation of c-ret expression by retinoic acid in rat metanephros: implication in nephron mass control.

Vitamin A and its derivatives have been shown to promote kidney development in vitro in a dose-dependent fashion. To address the molecular mechanisms by which all-trans-retinoic acid (RA) may regulate the nephron mass, rat kidneys were removed on embryonic day 14 (E14) and grown in organ culture under standard or RA-stimulated conditions. By using RT-PCR, we studied the expression of the glial cell line-derived neurotrophic factor (GDNF), its cell surface receptor-alpha (GDNFR-alpha), and the receptor tyrosine kinase c-ret, known to play a major role in renal organogenesis. Expression of GDNF and GDNFR-alpha transcripts was high at the time of explantation and remained unaffected in culture with or without RA. In contrast, c-ret mRNA level, which was low in E14 metanephros and dropped rapidly in vitro, was increased by RA in a dose-dependent manner. The same is true at the protein level. Exogenous GDNF barely promotes additional nephron formation in vitro. Thus the present data establish c-ret as a key target of retinoids during kidney organogenesis.

Localization of glial cell line-derived neurotrophic factor receptor alpha and c-ret mRNA in rat central nervous system.

Glial cell line-derived neurotrophic factor (GDNF) is a neurotrophic factor that influences the survival and function of several neuronal populations in the central (CNS) and peripheral nervous systems. The actions of GDNF are mediated by a multicomponent receptor complex composed of the tyrosine kinase product of c-ret and the ligand-binding protein GDNF receptor alpha (GDNFR-alpha). In the present study, we used in situ hybridization to localize cells expressing the mRNA for these GDNF receptor subunits in rat CNS. As reported previously, GDNFR-alpha and c-ret mRNA are present in the substantia nigra and ventral tegmental area, regions containing GDNF-responsive dopamine neurons. However, both mRNA were found in motor neurons of spinal cord and brainstem nuclei that innervate skeletal muscle. These areas include alpha motor neurons in the ventral horn of spinal cord and neurons in hypoglossal, facial, trigeminal, and abducens nuclei. In areas rostral to the substantia nigra, c-ret mRNA is not detected, whereas GDNFR-alpha is found in numerous brain structures, including the hippocampus, cortex, medial geniculate, and the medial habenula, the latter area expressing the highest levels of GDNFR-alpha mRNA in brain. These results provide evidence that c-ret and GDNFR-alpha mRNA are expressed in neuronal populations involved in motor function and provides further support for GDNF as a target-derived neurotrophic for these motor neurons. The observation that GDNFR-alpha mRNA is localized in several brain structures that do not contain detectable levels of c-ret mRNA indicates that either GDNFR-alpha utilizes signal transduction molecules other than c-ret in these areas or that other GDNF-like ligands that utilize GDNFR-alpha as a receptor may be present.

Developmental and denervation changes in c-ret proto-oncogene expression in chick motoneurons.

Receptor tyrosine kinases (RTKs) play important roles in cellular differentiation, survival, and proliferation. To search for genes involved in the survival of motoneurons, we isolated the RTKs specifically expressed on them. We used RT-PCR, by which RNA was obtained from purified embryonic day 5 (E5) chick motoneurons, and screened by in situ hybridization. Of 17 RTK cDNA clones, c-ret expression gradually increased in the motoneurons of the spinal cord during development. Compared with the expression of the neurotrophin receptor, TrkC, that of c-ret was very high in motoneurons from around E17 to adulthood. The level of c-ret expression on the motoneurons was unaffected by deafferentation at E2, but changed after denervation at post-hatching day 2. The in situ hybridization signal for c-ret mRNA increased slightly at day 1, then decreased progressively up to day 8, and increased again 2 weeks after sciatic nerve denervation in the motoneurons of the lumbar spinal cord. There were also changes in the signal of the lesioned sciatic nerve. In the most distal part from the lesioned site, the signal for c-ret mRNA significantly increased from day 3 on after denervation. These results suggested that the c-ret gene may encode the receptor for a factor involved in motoneuron differentiation and the promotion of regeneration of injured peripheral nerves.

Functional receptor for GDNF encoded by the c-ret proto-oncogene.

Glial-cell-line-derived neutrophic factor (GDNF) promotes the survival and phenotype of central dopaminergic noradrenergic and motor neurons, as well as various subpopulations of peripheral sensory and sympathetic neurons. GDNF is structurally related to members of the transforming growth factor (TGF)-beta superfamily, several members of which have well-characterized receptor systems; however, GDNF receptors still remain undefined. Here we show that GDNF binds to, and induces tyrosine phosphorylation of, the product of the c-ret proto-oncogene, an orphan receptor tyrosine kinase, in a GDNF-responsive motor-neuron cell line. Ret protein could also bind GDNF and mediate survival and growth responses to GDNF upon transfection into naive fibroblasts. Moreover, high levels of c-ret mRNA expression were found in dopaminergic neurons of the adult substantia nigra, where exogenous GDNF protected Ret-positive neurons from 6-hydroxydopamine-induced cell death. Thus the product of the c-ret proto-oncogene encodes a functional receptor for GDNF that may mediate its neurotrophic effects on motor and dopaminergic neurons.

Expression of the c-ret proto-oncogene during mouse embryogenesis

The c-ret proto-oncogene encodes a receptor tyrosine kinase whose normal function has yet to be determined. To begin to investigate the potential role of this gene in vertebrate development, we have isolated cDNA clones representing the murine c-ret gene, and have analyzed the pattern of expression during mouse embryogenesis, using northern blotting, in situ hybridization to histological sections and whole-mount hybridization histochemistry. c-ret transcripts were detected beginning at day 8.5 of embryogenesis, and were observed in a number of cell lineages in the developing peripheral and central nervous systems, as well as in the excretory system. In the cranial region at day 8.5-9.5, c-ret mRNA was restricted to a population of neural crest cells migrating from rhombomere 4 and forming the anlage of the facioacoustic ganglion, as well as to a closely associated domain of surface ectoderm and pharyngeal endoderm. At later stages (10.5-14.5 days), c-ret mRNA was observed in all cranial ganglia. In the peripheral nervous system of the trunk, c-ret was expressed in the autonomic ganglia and in subsets of cells in the dorsal root ganglia. In the enteric nervous system, c-ret was expressed in the presumptive enteric neuroblasts of the vagal crest (day 9.0-11.5), and in the myenteric ganglia of the gut (day 13.5-14.5). c-ret mRNA was observed in several regions of the central nervous system, including the undifferentiated neuroepithelial cells of the ventral neural tube (8.5 days), the motor neurons in the spinal cord and the hindbrain (10.5-14.5 days), the embryonic neuroretina (day 13.5) and the layers of the postnatal retina containing ganglion, amacrine and horizontal cells. Outside the nervous system, c-ret was expressed in the nephric (Wolffian) duct at day 8.5-10.5, the ureteric bud epithelium (but not the surrounding metanephric mesenchyme) at day 11.0-11.5, and the growing tips of the renal collecting ducts (but not the previously formed, subcortical portions of the collecting ducts, or the mesenchyme-derived renal vesicles) at day 13.5-17.5. Our results suggest that the c-ret gene may encode the receptor for a factor involved in the proliferation, migration, differentiation or survival of a variety of neuronal cell lineages, as well as in inductive interactions during organogenesis of the kidney.