Fig. 7

Methods for probing genotype–phenotype correlations. Genotype–phenotype correlations can be based on three different types of experimental setups, each with their own pros and cons. (Upper panel) In low throughput experiments, selected variants can be analyzed e.g. by animal studies, in primary cultures or by in vitro enzymatic and/or biophysical assays on purified protein. The results of such experiments are typically highly detailed and precise. However, determining variant effects in this manner is typically time consuming and expensive, and accordingly only a limited number of variants can be assessed in this manner. (Middel panel) Recent developments have allowed for high throughput measurements of variant effects. These so-called MAVEs typically probe either enzyme activity, protein abundance or protein–protein interactions. The advantages of these approaches are that they can inform on thousands of gene variants and depending on type of the assay, they can also provide mechanistic detail (e.g. reduced variant abundance indicates that the variant causes a reduced structural stability). However, in comparison with the low throughput analyses, the obtained results are less precise. (Lower panel) Computational predictors of variant effects typically rely on phylogenetic conservation or structural data. They are rapid and scalable to millions of variants. However, they do not typically provide mechanistic detail and are still imperfect predictors of pathogenicity. Figure compiled using Inkscape (v1.3). Parts of the figure were made using BioRender.com and PyMOL (v2.5.2) using the PDB entry 2O4H [44]