Combined genetic/epigenetic manipulations to promote CST axon regeneration
Embryonic neurons and peripheral neurons respond to axon injury with activation of transcriptional programs conducive to regrowth. In contrast, mature CNS neurons fail to reactivate expression of crucial regeneration associated genes (RAGs) in response to injury, resulting in failed axon growth and permanent loss of function. Forced reexpression of key RAGs in mature CNS neurons is a promising strategy for therapeutic intervention. We and others have shown that ectopic expression of pro-regenerative factors such as KLF7 and Sox11 promotes axon regeneration in CNS neurons. To build on this success, we are pursuing two strategies to further improve the growth response evoked by these TFs. First, we are testing the hypothesis that the effects may be enhanced by the presence of additional TFs that work synergistically to induce transcriptional programs supportive of growth. Indeed, research in the optic system has revealed that combinatorial genetic interventions are more efficient in improving regenerative outcomes. We are therefore systematically testing TF combinations that include established RAGs as well as novel targets discovered in our lab, in cell culture assays of neurite outgrowth and in vivo in a mouse model of spinal cord injury. Second, we are testing the hypothesis that TF efficacy can be improved by enhancing DNA accessibility in genomic loci corresponding to key RAGs in CNS neurons. Our preliminary data show that chromatin regions surrounding key RAGs progressively get restricted as neurons age, supporting the hypothesis that the developmental decline in regenerative ability may be accompanied by a corresponding decrease in RAG promoter accessibility for pro-regenerative TFs. We are currently combining proregenerative TFs along with treatments that relax chromatin in assays of axon outgrowth. Overall, this research is expected to reveal novel molecular regulatory codes that can be manipulated to improve axon regeneration after injury.
Ishwariya Venkatesh, Matthew Simpson, Zimei Wang, Ben Callif, Denise Coley, Murray Blackmore Department of Biomedical Sciences – Marquette University, Milwaukee