Norepinephrine transport-mediated gene expression in noradrenergic neurogenesis.
BACKGROUND
We have identified a differential gene expression profile in neural crest stem cells that is due to deletion of the norepinephrine transporter (NET) gene. NET is the target of psychotropic substances, such as tricyclic antidepressants and the drug of abuse, cocaine. NET mutations have been implicated in depression, anxiety, orthostatic intolerance and attention deficit hyperactivity disorder (ADHD). NET function in adult noradrenergic neurons of the peripheral and central nervous systems is to internalize norepinephrine from the synaptic cleft. By contrast, during embryogenesis norepinephrine (NE) transport promotes differentiation of neural crest stem cells and locus ceruleus progenitors into noradrenergic neurons, whereas NET inhibitors block noradrenergic differentiation. While the structure of NET und the regulation of NET function are well described, little is known about downstream target genes of norepinephrine (NE) transport.


RESULTS
We have prepared gene expression profiles of in vitro differentiating wild type and norepinephrine transporter-deficient (NETKO) mouse neural crest cells using long serial analysis of gene expression (LongSAGE). Comparison analyses have identified a number of important differentially expressed genes, including genes relevant to neural crest formation, noradrenergic neuron differentiation and the phenotype of NETKO mice. Examples of differentially expressed genes that affect noradrenergic cell differentiation include genes in the bone morphogenetic protein (BMP) signaling pathway, the Phox2b binding partner Tlx2, the ubiquitin ligase Praja2, and the inhibitor of Notch signaling, Numbl. Differentially expressed genes that are likely to contribute to the NETKO phenotype include dopamine-beta-hydroxylase (Dbh), tyrosine hydroxylase (Th), the peptide transmitter 'cocaine and amphetamine regulated transcript' (Cart), and the serotonin receptor subunit Htr3a. Real-time PCR confirmed differential expression of key genes not only in neural crest cells, but also in the adult superior cervical ganglion and locus ceruleus. In addition to known genes we have identified novel differentially expressed genes and thus provide a valuable database for future studies.


CONCLUSIONS
Loss of NET function during embryonic development in the mouse deregulates signaling pathways that are critically involved in neural crest formation and noradrenergic cell differentiation. The data further suggest deregulation of signaling pathways in the development and/or function of the NET-deficient peripheral, central and enteric nervous systems.
Hu
Yao Fei
YF
Department of Cell Biology, Neurobiology and Anatomy, Medical College of Wisconsin, Milwaukee, WI 53226, USA. yhu@mcw.edu
Caron
Marc G
MG
Sieber-Blum
Maya
M
D000818:Animals; D001692:Biological Transport; D055418:Bone Morphogenetic Protein 6 / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D005260:Female; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D015723:Gene Library; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry; D047908:Intracellular Signaling Peptides and Proteins; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D009432:Neural Crest / Q000166:cytology / Q000196:embryology / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D009638:Norepinephrine / Q000378:metabolism*; D050484:Norepinephrine Plasma Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D011985:Receptors, Serotonin / Q000235:genetics / Q000378:metabolism; D044406:Receptors, Serotonin, 5-HT3; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D000098882:Cocaine- and Amphetamine-Regulated Transcript Protein
C525817:Bmp6 protein, mouse; D055418:Bone Morphogenetic Protein 6; D018398:Homeodomain Proteins; C511230:Htr3a protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins; D050484:Norepinephrine Plasma Membrane Transport Proteins; C106292:Numbl protein, mouse; D011985:Receptors, Serotonin; D044406:Receptors, Serotonin, 5-HT3; C107836:Tlx2 protein, mouse; D000098882:Cocaine- and Amphetamine-Regulated Transcript Protein; D009638:Norepinephrine