Cytoskeletal Keratins Are Overexpressed in a Zebrafish Model 2 of Idiopathic Scoliosis

Idiopathic scoliosis (IS) is a three-dimensional rotation of the spine >10 degrees with an unknown etiology. Our laboratory established a late-onset IS model in zebrafish (Danio rerio) containing a deletion in kif7. 25% of kif7co63/co63 zebrafish develop spinal curvatures and are otherwise developmentally normal, although the molecular mechanisms underlying the scoliosis are unknown. To define transcripts associated with scoliosis in this model, we performed bulk mRNA sequencing on 6 weeks past fertilization (wpf) kif7co63/co63 zebrafish with and without scoliosis. Additionally, we sequenced kif7co63/co63, kif7co63/+, and AB zebrafish (n= 3 per genotype). Sequencing reads were aligned to the GRCz11 genome and FPKM values were calculated. Differences between groups were calculated for each transcript by t-test. Principal component analysis showed that transcriptomes clustered by sample age and genotype. kif7 mRNA was mildly reduced in both homozygous and heterozygous zebrafish compared to AB. Sonic hedgehog target genes were upregulated in kif7co63/co63 zebrafish over AB, but no difference was detected between scoliotic and non-scoliotic mutants. The top upregulated genes in scoliotic zebrafish were cytoskeletal keratins.. Pankeratin staining of 6 wpf scoliotic and non-scoliotic kif7co63/co63 zebrafish showed increased keratin levels within the zebrafish musculature and intervertebral disc (IVD). Keratins are major components of the embryonic notochord, and aberrant keratin expression has been associated with intervertebral disc degeneration (IVDD) in both zebrafish and humans. The role of increased keratin accumulation as a molecular mechanism associated with the onset of scoliosis warrants further study. Comparative gene expression profiling analysis of RNA-seq data in 6 week post fertilization (wpf) and 4 days post fertilization (dpf) zebrafish. Samples include 6wpf wild-type (n=3), 6wpf kif7co63/+ (n=3), 6wpf non-scoliotic kif7co63/co63 (n=3), 6wpf scoliotic kif7co63/co63, 4dpf wild type (n=2), 4dpf kif7co63/+ (n=2), and 4dpf kif7co63/co63 (n=2).