Identification of transcription factor combinations that promote axonal sprouting in the injured spinal cord and regeneration-associated gene expression

ABSTRACT: Axon regeneration after injury to the central nervous system is limited by an inhibitory environment but also because injured neurons fail to initiate expression of regeneration associated genes (RAGs), thought to be required for robust long-distance growth. The potential of strong RAG expression to promote CNS regeneration is exemplified by the conditioning lesion model, whereby peripheral nerve injury promotes regeneration of centrally projecting branches of the injured neurons. RAG expression could potentially be induced by delivery of the right combination of transcription factors (TF), but the specific TFs required remain to be identified. We here begin with a focused bioinformatics approach, analyzing binding site motifs in the promoters of the RAG program to identify nine candidate growth-promoting TFs. In vitro screening then identified two combinations, KLF7/MEF2 and ATF3/KLF7/MEF2, that had potent neurite-growth promoting activity, the latter being the more powerful. We expressed TFs in vivo in L4/L5 dorsal root ganglia to test whether these combinations would promote regeneration and activate the RAG program, mimicking the conditioning lesion effect. KLF7 and MEF2 were also tested individually for comparison. KLF7/MEF2, but not ATF3/KLF7/MEF2 or the individual TFs, promoted axonal sprouting into the lesion site and improved functional recovery. We also generated gene expression profiles of laser dissected dorsal root ganglion neurons specifically expressing these TF combinations, and of DRG neurons that had been axotomized. Unexpectedly, the MEF2-VP16 construct used had little transcriptional activity in vivo, suggesting additional steps may be required to achieve full MEF2 activity. All combinations except MEF2 alone induced RAG expression mirroring that induced by axotomy to significant extents, while ATF3/KLF7/MEF2, KLF7 and ATF3, but not KLF7/MEF2 also induced apoptosis-related genes which may hinder regeneration. In conclusion, the combination KLF7/MEF2 partially mimics the conditioning lesion effect and promotes functional improvement.

ORGANISM(S): Rattus norvegicus

PROVIDER: GSE270874 | GEO | 2025/02/08

REPOSITORIES: GEO