<p class="first" id="d1902798e144">Phosphatase and tensin homolog (
<i>PTEN</i>) is a tumor suppressor frequently mutated in diverse cancers. Germline
<i>PTEN</i> mutations are also associated with a range of clinical outcomes, including
PTEN hamartoma
tumor syndrome (PHTS) and autism spectrum disorder (ASD). To empower new insights
into PTEN function and clinically relevant genotype-phenotype relationships, we systematically
evaluated the effect of
<i>PTEN</i> mutations on lipid phosphatase activity
<i>in vivo</i>. Using a massively parallel approach that leverages an artificial humanized
yeast
model, we derived high-confidence estimates of functional impact for 7,244 single
amino acid PTEN variants (86% of possible). We identified 2,273 mutations with reduced
cellular lipid phosphatase activity, which includes 1,789 missense mutations. These
data recapitulated known functional findings but also uncovered new insights into
PTEN protein structure, biochemistry, and mutation tolerance. Several residues in
the catalytic pocket showed surprising mutational tolerance. We identified that the
solvent exposure of wild-type residues is a critical determinant of mutational tolerance.
Further, we created a comprehensive functional map by leveraging correlations between
amino acid substitutions to impute functional scores for all variants, including those
not present in the assay. Variant functional scores can reliably discriminate likely
pathogenic from benign alleles. Further, 32% of ClinVar unclassified missense variants
are phosphatase deficient in our assay, supporting their reclassification. ASD-associated
mutations generally had less severe fitness scores relative to PHTS-associated mutations
(p = 7.16 × 10
<sup>−5</sup>) and a higher fraction of hypomorphic mutations, arguing for continued
genotype-phenotype
studies in larger clinical datasets that can further leverage these rich functional
data.
</p>