Advertisement

pp 1-13 | Cite as

Usage of Bacterial Artificial Chromosomes for Studying BDNF Gene Regulation in Primary Cultures of Cortical Neurons and Astrocytes

  • Kaur Jaanson
  • Angela Pärn
  • Tõnis Timmusk
Protocol
Part of the Neuromethods book series

Abstract

BDNF gene has a complex structure and is regulated by multiple time- and stimulus-dependent promoters and distal regulatory elements. To better understand the interplay between these regulatory units, it is beneficial to study BDNF regulation in the context of a larger gene locus. The bacterial artificial chromosome (BAC) system allows propagating up to 300 kb long genomic regions, enabling to study the whole BDNF gene. Here, we present the methods for usage of BACs for studying BDNF gene regulation in primary cultures of rat cortical neurons and astrocytes.

Keywords

Astrocytes Bacterial artificial chromosome BDNF Neurons Primary culture Transfection 

References

  1. 1.
    Binder DK, Scharfman HE (2004) Brain-derived neurotrophic factor. Growth Factors Chur Switz 22(3):123–131Google Scholar
  2. 2.
    Huang EJ, Reichardt LF (2001) Neurotrophins: roles in neuronal development and function. Annu Rev Neurosci 24:677–736Google Scholar
  3. 3.
    Barde YA, Edgar D, Thoenen H (1982) Purification of a new neurotrophic factor from mammalian brain. EMBO J 1(5):549–553Google Scholar
  4. 4.
    Poo MM (2001) Neurotrophins as synaptic modulators. Nat Rev Neurosci 2(1):24–32Google Scholar
  5. 5.
    Bramham CR, Panja D (2014) BDNF regulation of synaptic structure, function, and plasticity. Neuropharmacology 76(Part C):601–602Google Scholar
  6. 6.
    Egan MF, Kojima M, Callicott JH, Goldberg TE, Kolachana BS, Bertolino A et al (2003) The BDNF val66met polymorphism affects activity-dependent secretion of BDNF and human memory and hippocampal function. Cell 112(2):257–269Google Scholar
  7. 7.
    Alsina B, Vu T, Cohen-Cory S (2001) Visualizing synapse formation in arborizing optic axons in vivo: dynamics and modulation by BDNF. Nat Neurosci 4(11):1093–1101Google Scholar
  8. 8.
    Zafra F, Lindholm D, Castrén E, Hartikka J, Thoenen H (1992) Regulation of brain-derived neurotrophic factor and nerve growth factor mRNA in primary cultures of hippocampal neurons and astrocytes. J Neurosci 12(12):4793–4799Google Scholar
  9. 9.
    Dougherty KD, Dreyfus CF, Black IB (2000) Brain-derived neurotrophic factor in astrocytes, oligodendrocytes, and microglia/macrophages after spinal cord injury. Neurobiol Dis 7(6 Pt B):574–585Google Scholar
  10. 10.
    Aid T, Kazantseva A, Piirsoo M, Palm K, Timmusk T (2007) Mouse and rat BDNF gene structure and expression revisited. J Neurosci Res 85(3):525–535Google Scholar
  11. 11.
    Metsis M, Timmusk T, Arenas E, Persson H (1993) Differential usage of multiple brain-derived neurotrophic factor promoters in the rat brain following neuronal activation. Proc Natl Acad Sci U S A 90(19):8802–8806Google Scholar
  12. 12.
    Timmusk T, Palm K, Metsis M, Reintam T, Paalme V, Saarma M et al (1993) Multiple promoters direct tissue-specific expression of the rat BDNF gene. Neuron 10(3):475–489Google Scholar
  13. 13.
    Liu Q-R, Lu L, Zhu X-G, Gong J-P, Shaham Y, Uhl GR (2006) Rodent BDNF genes, novel promoters, novel splice variants, and regulation by cocaine. Brain Res 1067(1):1–12Google Scholar
  14. 14.
    Liu Q-R, Walther D, Drgon T, Polesskaya O, Lesnick TG, Strain KJ et al (2005) Human brain derived neurotrophic factor (BDNF) genes, splicing patterns, and assessments of associations with substance abuse and Parkinson’s disease. Am J Med Genet B Neuropsychiatr Genet 134B(1):93–103Google Scholar
  15. 15.
    Pruunsild P, Kazantseva A, Aid T, Palm K, Timmusk T (2007) Dissecting the human BDNF locus: bidirectional transcription, complex splicing, and multiple promoters. Genomics 90(3):397–406Google Scholar
  16. 16.
    Pruunsild P, Sepp M, Orav E, Koppel I, Timmusk T (2011) Identification of cis-elements and transcription factors regulating neuronal activity-dependent transcription of human BDNF gene. J Neurosci 31(9):3295–3308Google Scholar
  17. 17.
    Koppel I, Timmusk T (2013) Differential regulation of Bdnf expression in cortical neurons by class-selective histone deacetylase inhibitors. Neuropharmacology 75C:106–115Google Scholar
  18. 18.
    Tuvikene J, Pruunsild P, Orav E, Esvald E-E, Timmusk T (2016) AP-1 transcription factors mediate BDNF-positive feedback loop in cortical neurons. J Neurosci 36(4):1290–1305Google Scholar
  19. 19.
    Hosoda F, Nishimura S, Uchida H, Ohki M (1990) An F factor based cloning system for large DNA fragments. Nucleic Acids Res 18(13):3863–3869Google Scholar
  20. 20.
    Shizuya H, Kouros-Mehr H (2001) The development and applications of the bacterial artificial chromosome cloning system. Keio J Med 50(1):26–30Google Scholar
  21. 21.
    Shizuya H, Birren B, Kim UJ, Mancino V, Slepak T, Tachiiri Y et al (1992) Cloning and stable maintenance of 300-kilobase-pair fragments of human DNA in Escherichia coli using an F-factor-based vector. Proc Natl Acad Sci U S A 89(18):8794–8797Google Scholar
  22. 22.
    Muyrers JP, Zhang Y, Testa G, Stewart AF (1999) Rapid modification of bacterial artificial chromosomes by ET-recombination. Nucleic Acids Res 27(6):1555–1557Google Scholar
  23. 23.
    Yang XW, Model P, Heintz N (1997) Homologous recombination based modification in Escherichia coli and germline transmission in transgenic mice of a bacterial artificial chromosome. Nat Biotechnol 15(9):859–865Google Scholar
  24. 24.
    Shen W, Huang Y, Tang Y, Liu D-P, Liang C-C (2005) A general method to modify BACs to generate large recombinant DNA fragments. Mol Biotechnol 31(3):181–186Google Scholar
  25. 25.
    Bird AW, Erler A, Fu J, Hériché J-K, Maresca M, Zhang Y et al (2012) High-efficiency counterselection recombineering for site-directed mutagenesis in bacterial artificial chromosomes. Nat Methods 9(1):103–109Google Scholar
  26. 26.
    Hartwich H, Nothwang H (2012) An easy and versatile 2-step protocol for targeted modification and subcloning of DNA from bacterial artificial chromosomes using non-commercial plasmids. BMC Res Notes 5(1):156Google Scholar
  27. 27.
    Gong S, Zheng C, Doughty ML, Losos K, Didkovsky N, Schambra UB et al (2003) A gene expression atlas of the central nervous system based on bacterial artificial chromosomes. Nature 425(6961):917–925Google Scholar
  28. 28.
    Illenye S, Heintz NH (2004) Functional analysis of bacterial artificial chromosomes in mammalian cells: mouse Cdc6 is associated with the mitotic spindle apparatus. Genomics 83(1):66–75Google Scholar
  29. 29.
    Montigny WJ, Houchens CR, Illenye S, Gilbert J, Coonrod E, Chang YC et al (2001) Condensation by DNA looping facilitates transfer of large DNA molecules into mammalian cells. Nucleic Acids Res 29(9):1982–1988Google Scholar
  30. 30.
    Montigny WJ, Phelps SF, Illenye S, Heintz NH (2003) Parameters influencing high-efficiency transfection of bacterial artificial chromosomes into cultured mammalian cells. Biotechniques 35(4):796–807Google Scholar
  31. 31.
    Schwank G, Andersson-Rolf A, Koo B-K, Sasaki N, Clevers H (2013) Generation of BAC transgenic epithelial organoids. PLoS One 8(10):e76871Google Scholar
  32. 32.
    Mader A, Prewein B, Zboray K, Casanova E, Kunert R (2013) Exploration of BAC versus plasmid expression vectors in recombinant CHO cells. Appl Microbiol Biotechnol 97(9):4049–4054Google Scholar
  33. 33.
    Kunert R, Casanova E (2013) Recent advances in recombinant protein production: BAC-based expression vectors, the bigger the better. Bioengineered 4(4):258–261Google Scholar
  34. 34.
    Smith-Hicks C, Xiao B, Deng R, Ji Y, Zhao X, Shepherd JD et al (2010) SRF binding to SRE 6.9 in the Arc promoter is essential for LTD in cultured Purkinje cells. Nat Neurosci 13(9):1082–1089Google Scholar
  35. 35.
    Koppel I, Aid-Pavlidis T, Jaanson K, Sepp M, Pruunsild P, Palm K et al (2009) Tissue-specific and neural activity-regulated expression of human BDNF gene in BAC transgenic mice. BMC Neurosci 10:68Google Scholar
  36. 36.
    Koppel I, Aid-Pavlidis T, Jaanson K, Sepp M, Palm K, Timmusk T (2010) BAC transgenic mice reveal distal cis-regulatory elements governing BDNF gene expression. Genesis 48(4):214–219Google Scholar
  37. 37.
    Jaanson K, Sepp M, Aid-Pavlidis T, Timmusk T (2014) BAC-based cellular model for screening regulators of BDNF gene transcription. BMC Neurosci 15:75Google Scholar
  38. 38.
    Koppel I, Jaanson K, Klasche A, Tuvikene J, Tiirik T, Pärn A et al (2018) Dopamine cross-reacts with adrenoreceptors in cortical astrocytes to induce BDNF expression, CREB signaling and morphological transformation. Glia 66(1):206–216Google Scholar

Copyright information

© Springer Science+Business Media New York 2018

Authors and Affiliations

  1. 1.Department of Chemistry and BiotechnologyTallinn University of TechnologyTallinnEstonia

Personalised recommendations