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BMC Neurosci. 2014 Jun 18;15:75. doi: 10.1186/1471-2202-15-75.

BAC-based cellular model for screening regulators of BDNF gene transcription.

Author information

1
Department of Gene Technology, Tallinn University of Technology, Akadeemia tee 15, 12618 Tallinn, Estonia. kaur.jaanson@ttu.ee.

Abstract

BACKGROUND:

Brain derived neurotrophic factor (BDNF) belongs to a family of structurally related proteins called neurotrophins that have been shown to regulate survival and growth of neurons in the developing central and peripheral nervous system and also to take part in synaptic plasticity related processes in adulthood. Since BDNF is associated with several nervous system disorders it would be beneficial to have cellular reporter system for studying its expression regulation.

METHODS:

Using modified bacterial artificial chromosome (BAC), we generated several transgenic cell lines expressing humanised Renilla luciferase (hRluc)-EGFP fusion reporter gene under the control of rat BDNF gene regulatory sequences (rBDNF-hRluc-EGFP) in HeLa background. To see if the hRluc-EGFP reporter was regulated in response to known regulators of BDNF expression we treated cell lines with substances known to regulate BDNF and also overexpressed transcription factors known to regulate BDNF gene in established cell lines.

RESULTS:

rBDNF-hRluc-EGFP cell lines had high transgene copy numbers when assayed with qPCR and FISH analysis showed that transgene was maintained episomally in all cell lines. Luciferase activity in transgenic cell lines was induced in response to ionomycin-mediated rise of intracellular calcium levels, treatment with HDAC inhibitors and by over-expression of transcription factors known to increase BDNF expression, indicating that transcription of the transgenic reporter is regulated similarly to the endogenous BDNF gene.

CONCLUSIONS:

Generated rBDNF-hRluc-EGFP BAC cell lines respond to known modulators of BDNF expression and could be used for screening of compounds/small molecules or transcription factors altering BDNF expression.

PMID:
24943717
PMCID:
PMC4071165
DOI:
10.1186/1471-2202-15-75
[Indexed for MEDLINE]
Free PMC Article
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