Glial-specific gene expression after contusive spinal cord injury

University Of Louisville, Louisville KY

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Abstract

No clinically applicable treatments exist to improve functional outcome after spinal cord injury (SCI). In part, such a lack of progress is due to poor understanding of the complex pathology of this injury. The SCI response is cell type-specific and evolves with time post-injury. Acutely after injury, oligodendrocytes (OL) display disruption of proteostasis including endoplasmic reticulum stress (ERSR) and integrated stress (ISR) responses. The ISR leads to apoptosis and white matter loss that underlies many deficits of sensation and locomotion. Later, oligodendrocyte precursor cells (OPCs) proliferate in an attempt to repair lost myelin. However, that response is limited by inhibitory signals that block OPC differentiation. While global spinal cord transcriptomics have enabled a systematic knowledge about the SCI response, conclusions from these efforts are limited by technological barriers and the complexity of spinal cord anatomy. Thus, SCI-associated changes in mRNA levels may not translate into parallel effects on protein expression. Moreover, homogenized fragments of whole spinal cord used for traditional transcriptomic experiments make identification of novel components of the cell-specific injury response challenging. These limitations may be overcome by Translating Ribosome Affinity Purification (TRAP) technology which isolates and analyzes only mRNAs that are associated with ribosomes (i.e. being likely translated) from specific cell populations that were marked genetically with a ribosomal tag. Thus, we propose to apply TRAP to test the hypothesis that after SCI, both transcriptional and translational reprogramming regulate the expression of critical components of the injury response in a cell type- specific manner. Furthermore, translational regulation will be of particular significance to launch proteostasis responses in OPCs and OLs. We also propose that in the context of OPC/OLs that respond to SCI, identifying translated mRNAs for transcription factors will uncover new targets that can be targeted for OL protection and/or remyelination. Focusing on the injury epicenter, we will characterize translated transcriptome profiles of OPC/OLs at 2, 10 and 42 days after moderate mouse contusive SCI inOPC/OLs from transgenic mice that express EGFP-L10 ribosome tag in a Cre dependent manner selectively in these cells (specific aim 1). In addition, we will use the resulting data sets to identify transcription factors (TF) that orchestrate OPC/OL responses to SCI (specific aim 2). We expect to characterize SCI-associated gene expression events in OPC/OLs with unprecedented accuracy. By focusing on ribosome-associated transcripts, our data will paint a landscape of complete gene expression events that reach the protein level. Such a landscape is unavailable for SCI-challenged OPC/OLs on a whole genome scale. Therefore, this high risk/high reward proposal may redefine the SCI response of OPC/OLs and identify novel targets for white matter protection and/or remyelination therapies.