Multiple mechanisms lead to differential loss-of-function effects of pathogenic SARS2 variants: implications for genotype-phenotype analyses

Seryl-tRNA synthetase 2 (SARS2) encodes the enzyme responsible for charging tRNA with serine in the mitochondria. SARS2 has been associated with a spectrum of recessive diseases including HUPRA syndrome and progressive spastic paresis (PSP). Previous studies showed that pathogenic SARS2 variants cause decreased tRNA charging; however, the mechanism by which specific variants lead to distinct recessive phenotypes has not been defined. To address this lack of knowledge, we studied an allelic series of pathogenic SARS2 variants for differential effects on mitochondrial function. We generated a SARS2 knockout Hap1 cell line with a second, doxycycline-inducible wild-type copy of SARS2 to support cell growth. We first assessed the effects of ablated SARS2 expression on the transcriptome and on mitochondrial function. We found that, in the absence of SARS2 expression, transcriptional changes indicate activation of cellular stress responses including the integrated stress response. Furthermore, loss of SARS2 resulted in severely depleted oxygen consumption in Seahorse assays. We next individually transduced an allelic series of 11 pathogenic SARS2 variants into this cell model to assess the effects of each variant on mitochondrial function. These efforts revealed compelling variant-dependent effects on oxygen consumption that will be useful for phenotype-genotype correlations. Interestingly, the most commonly detected pathogenic SARS2 variant (R402H) had no effect on mitochondrial function in our model system. We performed computational and functional studies, which showed that this missense variant has a partial effect on exon inclusion, which predicts a hypomorphic allele consistent with certain patients being reported as homozygous for this variant. Overall, this study expands our understanding of SARS2 biology, reveals the differential effects that pathogenic variants have on SARS2 function, and provides the foundation for defining the clinical heterogeneity of patient phenotypes.