Learning-Dependent Transcriptional Regulation of BDNF by its Truncated Protein Isoform in Turtle

Abstract

The vertebrate brain-derived neurotrophic factor (BDNF) gene produces a number of alternatively spliced transcripts only some of which generate the BDNF protein required for synaptic plasticity and learning. Many of the transcripts are uncharacterized and are of unknown biological significance. Previously, we described alternative splicing within the protein-coding sequence of the BDNF gene in the pond turtle (tBDNF) that generates a functionally distinct truncated protein isoform (trcBDNF) that is regulated during a neural correlate of eyeblink classical conditioning in ex vivo brainstem preparations. We hypothesized that trcBDNF has a dominant negative function because of its anticorrelated expression pattern compared to its full-length BDNF counterpart. The data presented here suggests that trcBDNF functions as a transcriptional repressor of a conditioning-inducible downstream tBDNF promoter that controls expression of full-length BDNF required for learning. First, expression of full-length transcripts is negatively correlated with trcBDNF; transcripts are inhibited when endogenous trcBDNF is ectopically induced and expressed when trcBDNF is inhibited. Second, ChIP-qPCR assays of a recombinant trcBDNF protein, RtrcBDNF, show strong binding to the downstream tBDNF exon III promoter that corresponds with inhibition of conditioning. Third, deletions of the C-terminus of RtrcBDNF result in inhibition of promoter binding and conditioning acquisition when a tropomyosin receptor kinase B (TrkB) binding site is accounted for. Finally, microinjection of RtrcBDNF directly into brainstem preparations inhibits conditioning. These data reveal a new mechanism of activity-dependent BDNF transcriptional regulation and suggest that BDNF is an autoregulatory gene. How generalizable this mechanism is across plasticity genes remains to be elucidated.