Mendelian gene identification through mouse embryo viability screening

Abstract

The diagnostic rate of Mendelian disorders in sequencing studies continues to increase, along with the pace of novel disease gene discovery. However, variant interpretation in novel genes not currently associated with disease is particularly challenging and strategies combining gene functional evidence with approaches that evaluate the phenotypic similarities between patients and model organisms have proven successful. A full spectrum of intolerance to loss-of-function variation has been previously described, providing evidence that gene essentiality should not be considered as a simple and fixed binary property. Here we further dissected this spectrum by assessing the embryonic stage at which homozygous loss-of-function results in lethality in mice from the International Mouse Phenotyping Consortium, classifying the set of lethal genes into one of three windows of lethality: early, mid or late gestation lethal. We studied the correlation between these windows of lethality and various gene features including expression across development, paralogy and constraint metrics together with human disease phenotypes, and found that the members of the early gestation lethal category show distinctive characteristics and a strong enrichment for genes linked with recessive forms of inherited metabolic disease. Based on these findings, we explored a gene similarity approach for novel gene discovery focused on this subset of lethal genes. Finally, we investigated unsolved cases from the 100,000 Genomes Project recruited under this disease category to look for signs of enrichment of biallelic predicted pathogenic variants among early gestation lethal genes and highlight two novel candidates with phenotypic overlap between the patients and the mouse knockout.

Lethal phenotypes in Mendelian disorders

“…Robbins et al suggested that lethality occurring early in the pregnancy would involve genes related to fundamental cellular processes, while genes associated to developmental biological processes would be associated to death occurring at a later fetal developmental stage 39 . The analysis of mouse knockout lethal genes confirms the association between the embryonic stage at which lethality occurs and the different biological processes ascribed to each corresponding genes 5,30 .…”

“…fetal hydrops, a unique severe often lethal prenatal phenotype can be associated to cardiovascular, immunological, or the haematological category. More granular associations can also be found when we explore the exact embryonic stage at which the mouse embryo dies 30 .…”

“…When we break down the set of mouse lethal genes into more granular categories, i.e. cellular lethal and developmental lethal genes 5 , or early gestation, mid gestation and late gestation lethal genes 30 , we observe that this enrichment is not uniform, and that the sets of developmental lethal or late gestation lethal genes are the ones more significantly enriched in human disease genes. This observation led to the hypothesis that the set of cellular or early gestation lethal genes may be associated with early embryonic lethality in humans, a phenotype not well captured in current gene-disease association databases and difficult to ascertain, as it will be explained in the next section.…”

“…Overall, the set of genes that are lethal in the mouse and not currently associated with human disease constitutes a powerful source of genes potentially linked to severe, multisystemic, early onset disorders in humans, including those with prenatal and neonatal lethal phenotypes, e.g. families with a history of miscarriages followed by a child with a severe, early age of onset condition of suspected monogenic origin 5,26,30,32 . Gene and variant prioritisation strategies leveraging this information have been successful in identifying novel neurodevelopmental disease genes 33,34 .…”

Impact of Essential Genes on the Success of Genome Editing Experiments Generating 3,313 New Genetically Engineered Mouse Lines

“…The effect of cell essentiality or lethality can apply to generating lines with disease-associated variants such as single nucleotide variants. Given the high proportion of essential genes among human disease genes 19,27 , generating mouse models of human disease may require preserving one copy of the wild-type allele, that is producing heterozygous founders or conditional variant alleles, to establish a new mouse line. Overall, the use of Cas9 is a robust and flexible method to generate gene-edited mouse lines.…”