NFAT5/TonEBP controls early acquisition of notochord phenotypic markers, collagen composition, and sonic hedgehog signaling during mouse intervertebral disc embryogenesis.

High osmolarity, bound water, and hydrostatic pressure contribute to notochord mechanics and its morphogenesis into the nucleus pulposus (NP) compartment of the intervertebral disc. Indeed, the osmoadaptive transcription factor, nuclear factor of activated T-cells 5 (NFAT5 aka TonEBP), is robustly expressed by resident cells of the notochord and NP. Nevertheless, the molecular mechanisms that drive notochord osmoregulation and the functions of NFAT5 in disc embryogenesis remain largely unexplored. In this study, we show that deletion of NFAT5 in mice results in delayed vertebral column development and a reduced NP aspect ratio in the caudal spine. This phenotype is associated with lower levels of the T-box transcription factor, Brachyury, delayed expression of notochord phenotypic markers, and decreased collagen II deposition in the perinotochordal sheath and condensing mesenchyme. In addition, NFAT5 mutants showed a stage-dependent dysregulation of sonic hedgehog (Shh) signaling with non-classical expression of Gli1. Generation of mice with notochord-specific deletion of IFT88 (ShhcreERT2;Ift88f/f) supported this mode of Gli1 regulation. Using isolated primary NP cells and bioinformatics approaches, we further show that Ptch1 and Smo expression is controlled by NFAT5 in a cell autonomous manner. Altogether, our results demonstrate that NFAT5 contributes to notochord and disc embryogenesis through its regulation of hallmark notochord phenotypic markers, extracellular matrix, and Shh signaling.