MicroRNA-1-associated effects of neuron-specific brain-derived neurotrophic factor gene deletion in dorsal root ganglia

Mol Cell Neurosci. 2016 Sep:75:36-43. doi: 10.1016/j.mcn.2016.06.003. Epub 2016 Jun 21.

Abstract

Background: MicroRNAs (miRNAs) regulate gene expression in physiological as well as in pathological processes, including chronic pain. Whether deletion of a gene can affect expression of the miRNAs that associate with the deleted gene mRNA remains elusive. We investigated the effects of brain-derived neurotrophic factor (Bdnf) gene deletion on the expression of miR-1 in dorsal root ganglion (DRG) neurons and its pain-associated downstream targets heat shock protein 60 (Hsp60) and connexin 43 (Cx43) in tamoxifen-inducible conditional knockout mice, Bdnf(fl/fl); Advillin-CreER(T2) (Bdnf cKO).

Results: Efficient Bdnf gene deletion was confirmed in DRG of Bdnf cKO mice by Real-Time qRT-PCR and ELISA 10days after completed tamoxifen treatment. In DRG, miR-1 expression was reduced 0.44-fold (p<0.05; Real-time qRT-PCR) in Bdnf cKO compared to floxed wildtype littermate control Bdnf(fl/fl) mice (WT). While Hsp60 protein expression was increased 1.85-fold (p<0.05; Western blot analysis), expression levels of Cx43 and the miR-1-associated transcription factors MEF2a and SRF remained unchanged. When analyzing Bdnf cKO mice 32days after complete tamoxifen treatment to investigate whether observed expression alterations remain permanently, we found no significant differences between Bdnf cKO and WT mice. However, miRNA microarray analysis revealed that 167 miRNAs altered (p<0.05) in DRG of these mice following Bdnf gene deletion.

Conclusions: Our results indicate that deletion of Bdnf in DRG neurons leads to a temporary dysregulation of miR-1, suggesting an impairment of a presumable feedback loop between BDNF protein and its targeting miR-1. This appears to affect its downstream protein Hsp60 and as a consequence might influence the phenotype after inducible Bdnf gene deletion. While this appears to be a MEF2a-/SRF-independent and transient effect, expression levels of various other miRNAs may remain permanently altered.

Keywords: BDNF-Advillin-Cre-ERT2; Bdnf; Dorsal root ganglion; Gene deletion; Neuropathic pain; miR-1; microRNA.

MeSH terms

  • Animals
  • Brain-Derived Neurotrophic Factor / genetics*
  • Brain-Derived Neurotrophic Factor / metabolism
  • Chaperonin 60 / genetics
  • Chaperonin 60 / metabolism
  • Connexin 43 / genetics
  • Connexin 43 / metabolism
  • Female
  • Ganglia, Spinal / metabolism*
  • Gene Deletion
  • Male
  • Mice
  • Mice, Inbred C57BL
  • MicroRNAs / genetics*
  • MicroRNAs / metabolism
  • Mitochondrial Proteins / genetics
  • Mitochondrial Proteins / metabolism

Substances

  • Brain-Derived Neurotrophic Factor
  • Chaperonin 60
  • Connexin 43
  • Hspd1 protein, mouse
  • MicroRNAs
  • Mirn1 microRNA, mouse
  • Mitochondrial Proteins