Transcriptional Activation of the Human Brain-derived Neurotrophic Factor Gene Promoter III by Dopamine Signaling in NT2/N Neurons

Abstract

Brain-derived neurotrophic factor (BDNF) 1 is a member of neurotrophin family, structurally related to nerve growth factor, neurotrophin-3, and neurotrophin-4/5. Its biological activity is mediated by tyrosine kinase receptor B (TrkB) and its downstream signaling (1). The gene is highly expressed and widely distributed in the central nervous system, and it plays a significant role in the maintenance of function and survival of neurons (for review, see Ref.2). Evidence accumulated in recent years suggests that BDNF is also involved in the modulation of synaptic activity in the adult brain, producing long lasting changes in synaptic structure and function (for reviews, see Refs. 3-5).There is a growing interest in BDNF as a potential therapeutic agent for neurodegenerative diseases, because its deficiency was found in brains of both Alzheimer's and Parkinson's patients (6 -10). Indeed, BDNF treatment has been shown not only to potentiate synaptic transmission in vivo (11, 12) but also to increase neuronal survival and augment some behavioral changes in animal models (13,14). However, more recent data indicate that BDNF can also induce behavioral sensitization by causing an overexpression of dopamine D3 receptors and could, actually, contribute to the amplification of pathophysiologies associated with conditions such as epilepsy, drug addiction, schizophrenia, and Parkinson's disease (15, 16). Clearly, further work is required to resolve some of these potential side-effects. In contrast to a large body of work on the temporal and spatial patterns of BDNF expression in neurodevelopment and neurodegeneration, relatively little is known about the transcriptional regulation of the human BDNF gene. This is partially due to the fact that the genomic structure of the human gene has not yet been fully elucidated. The gene was first localized to chromosome11p13 and predicted to consist of multiple exons (17), but the existence of multiple transcripts, derived from different exons, was demonstrated only recently by Aoyama et al. (18) in human neuroblastoma cells. A more detailed transcript mapping of an 810-kb region of chromosome 11p13-14 further defined its genomic localization, although no additional information on the actual structure of the gene itself was presented (19).The existence of multiple human BDNF transcripts (18) is consistent with a genomic structure similar to that of the rat gene, which consists of four short 5Ј exons, each controlled by a distinct promoter, and one 3Ј exon encoding the mature BDNF protein (20,21). In the rat, the four promoters direct expression of the BDNF gene in a tissue-specific manner, i.e. promoters I and II are active preferentially in neurons, whereas promoters III and IV are active both in neurons and in a limited number of non-neuronal tissues such as lung and heart (20, 21). Thus far, only a limited characterization of a 3.2-kb genomic fragment of the human gene containing some structural elements of a promoter was reported (22).Recently, two transcription factors, CREB and ...

Reversal of Human Cytomegalovirus Major Immediate-Early Enhancer/Promoter Silencing in Quiescently Infected Cells via the Cyclic AMP Signaling Pathway

“…The primary antibodies were used at 1:1,000 to 1:2,000 dilutions. They include anti-CREB [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24] , an immunoaffinity-purified polyclonal rabbit antibody raised against amino-terminal residues 5 to 24 of human CREB (Upstate Biotechnology, Lake Placid, NY); anti-CREB 260-280 , a monoclonal rabbit antibody raised against residues 260 to 280 (Epitomics, Burlingame, CA); anti-pCREB 122-136 , an affinity-purified polyclonal rabbit antibody raised against a synthetic phosphopeptide corresponding to residues 126 to 136 (phospho-Ser133) of rat CREB (Upstate Biotechnology); and anti-ATF-1 (25C10G), a monoclonal murine antibody raised against residues 39 to 271 of human ATF-1 (Santa Cruz Biotechnology, Inc., Santa Cruz, CA). HCMV IE1 p72 and IE2 p86 were detected using monoclonal murine antibody MAB810 (Chemicon International), which reacts to an epitope in both proteins.…”

“…The anti-pCREB 122-136 (␣-pCREB 122-136 ) cross-reacts with phospho-Ser63 in corresponding residues of ATF-1. The blot was reprobed with anti-CREB 5-24 (␣-CREB [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24] ) and again reprobed for ␤-tubulin. Autoradiographs were subjected to long or short exposures.…”

HBZ Stimulates Brain-Derived Neurotrophic Factor/TrkB Autocrine/Paracrine Signaling To Promote Survival of Human T-Cell Leukemia Virus Type 1-Infected T Cells

“…However, the BDNF-IV promoter proved to be unresponsive to HBZ in the reported assays. It is possible that this promoter does not encompass cis-elements affected by the viral protein, as the BDNF gene contains nine separate transcription start sites distributed over more than 60 kb of DNA (29,30,58,59). Alternatively, HBZ may activate transcription through a distal enhancer.…”

S/MAR-binding properties of Sox2 and its involvement in apoptosis of human NT2 neural precursors

“…40 At the end of the OGD treatment (designated 0 h), cells were removed from the chamber and returned to the incubator for 3, 4 or 24 h. Cell viability was assessed by the Trypan Blue exclusion assay. Labeled cells were counted using a hemocytometer.…”

“…Trace DNA contamination was removed using the DNA-free kit (Ambion Inc., Austin, TX, USA). Total RNA was reverse transcribed and cDNA was purified and quantified as described by Fang et al 40 A 220 bp cDNA fragment of Sox2 was amplified from 200 ng of template DNA using high-fidelity Taq polymerase (Clontech, Palo Alto, CA, USA) with the following primers: Sox2F: 5 0 ATGTACAAC ATGAT GGAGA CG 3 0 and SOX2R: 5 0 GCGCTTGCTGAT CTCCGAGT 3 0 . PCR conditions were: one cycle at 941C for 5 min, 30 cycles for 1 min at 941C, 1 min at 601C and 1 min at 721C, and one cycle at 721C for 5 min.…”