ABSTRACT: OSGEP gene encodes a KEOPS (Kinase, Endopeptidase and Other Proteins of Small size) complex subunit catalyzing N6-threonylcarbamoyladenosine (t6A) modification to A37 of ANN tRNAs. This modification enhances translation accuracy and efficiency. OSGEP mutations are linked to intellectual disability, developmental delay, seizures, microcephaly, other brain abnormalities and kidney problems together termed Galloway-Mowat Syndrome. KEOPS may also function in telomere length regulation, genome maintenance and DNA damage response (DDR). OSGEP knockdown in human cells impairs translation and cellular proliferation, increases ER stress and apoptosis, activates DDR and disrupts actin regulation. Frameshift zebrafish osgep mutants exhibit microcephaly, increased brain apoptosis, and early lethality. Mouse Osgep knockouts had significantly shorter cortex lengths, widths and cortex–midbrain midline lengths. A renal phenotype was not observed in either knockout model, possibly due to early lethality masking renal involvement in older animals. Based on human-genetic data, we generated a zebrafish osgep G177A knock-in mutant. osgepG177A/G177A larvae have smaller eyes and heads relative to wildtype. The mutants have growth defects and about 70% of them die during the first 4-5 weeks post-fertilization. We determined that the p53 pathway is activated in osgepG177A/G177A embryos and elevates apoptosis levels. However, loss of tp53 does not rescue the phenotypes or survival of osgep mutants. RNA-sequencing of single wildtype and osgepG177A/G177A larvae identified >1000 differentially-expressed genes. Up-regulated genes were associated with protein folding, endoplasmic reticulum, RNA processing/maturation and ribosome biogenesis. Down-regulated genes were strongly enriched among translation-related, developmental, immune and hematopoietic terms, inflammatory response as well as Ribosome, Proteasome, Ferroptosis, Apoptosis and Oxidative Phosphorylation pathways. Maternal-zygotic (MZ) osgepG177A/G177A embryos generated from the surviving zygotic mutants exhibit dramatically exacerbated phenotypes with dramatically increased activations of p53 pathway and various Unfolded Protein Response (UPR) genes. Loss of tp53 in osgepG177A/G177A MZ embryos further worsened their phenotypes and did not alter UPR gene up-regulation.
