Cytoskeletal Keratins Are Overexpressed in a Zebrafish Model 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. A total of 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 the 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.

MIRO2 Regulates Prostate Cancer Cell Growth via GCN1-Dependent Stress Signaling.

There is a continued need to identify novel therapeutic targets to prevent the mortality associated with prostate cancer. In this context, mitochondrial Rho GTPase 2 (MIRO2) mRNA was upregulated in metastatic prostate cancer compared with localized tumors, and higher MIRO2 levels were correlated with poor patient survival. Using human cell lines that represent androgen-independent or -sensitive prostate cancer, we showed that MIRO2 depletion impaired cell growth, colony formation, and tumor growth in mice. Network analysis of MIRO2's binding partners identified metabolism and cellular responses to extracellular stimuli as top overrepresented pathways. The top hit on our screen, General Control Nonderepressible 1 (GCN1), was overexpressed in prostate cancer, and interacted with MIRO2 in prostate cancer cell lines and in primary prostate cancer cells. Functional analysis of MIRO2 mutations present in patients with prostate cancer led to the identification of MIRO2 159L, which increased GCN1 binding. Importantly, MIRO2 was necessary for efficient GCN1-mediated GCN2 kinase signaling and induction of the transcription factor activating transcription factor 4 (ATF4) levels. Further, MIRO2's effect on regulating prostate cancer cell growth was mediated by ATF4. Finally, levels of activated GCN2 and ATF4 were correlated with MIRO2 expression in prostate cancer xenografts. Both MIRO2 and activated GCN2 levels were higher in hypoxic areas of prostate cancer xenografts. Overall, we propose that targeting the MIRO2-GCN1 axis may be a valuable strategy to halt prostate cancer growth. IMPLICATIONS: MIRO2/GCN1/GCN2 constitute a novel mitochondrial signaling pathway that controls androgen-independent and androgen-sensitive prostate cancer cell growth.