TNF-α Promotes Nuclear Enrichment of TonEBP/NFAT5 to Selectively Control Inflammatory but not Osmoregulatory Responses in Nucleus Pulposus Cells.

Intervertebral disc degeneration (IDD) causes chronic back pain and is linked to production of proinflammatory molecules by nucleus pulposus (NP) and other disc cells. Activation of TonicityResponsive Enhancer-Binding Protein (TonEBP)/NFAT5 by non-osmotic stimuli, including pro-inflammatory molecules, occurs in cells involved in immune response. However, whether inflammatory stimuli activate TonEBP in NP cells and if TonEBP controls inflammation during IDD is unknown. We show that TNF-α, but not IL-1β or LPS, promoted nuclear enrichment of TonEBP protein. However, TNF-α-mediated activation of TonEBP did not cause induction of osmo-regulatory genes. RNA-sequencing showed that 8.5% of TNF-α transcriptional responses were TonEBP-dependent and identified genes regulated by both TNF-α and TonEBP. These genes were over-enriched in pathways and diseases related to Inflammatory Response and Inhibition of Matrix Metalloproteases. Based on RNA-seq results, we further investigated regulation of novel TonEBP targets CXCL1, CXCL2, and CXCL3. TonEBP acted synergistically with TNF-α and LPS to induce CXCL1 proximal promoter activity. Interestingly, this regulation required a highly conserved NF-κB binding site but not a predicted TonE, suggesting crosstalk between these two members of the Rel family. Finally, analysis of human NP tissue showed that TonEBP expression correlated with canonical osmo-regulatory targets TauT/SLC6A6, SMIT/SLC5A3, and AR/AKR1B1, supporting in vitro findings that the inflammatory milieu during IDD does not interfere with TonEBP osmoregulation. In summary, while TonEBP participates in the pro-inflammatory response to TNF-α, therapeutic strategies targeting this transcription factor for treatment of disc disease must spare osmo-protective, pro-survival, and matrix homeostatic activities. [Abstract copyright: Copyright © 2017, The American Society for Biochemistry and Molecular Biology.]