Reduced serine synthesis in patient-derived retinal pigmented epithelium leads to localized lipid dysfunction in MacTel

Abstract

Purpose : Macular telangiectasia type 2 (MacTel) is an orphan macular degenerative disease with polygenic inheritance and variable penetrance. GWAS analyses and metabalomics screens have indicated a defect in serine metabolism in MacTel, and recent whole exome sequencing has identified coding variants in PHGDH linked to MacTel in a subset of patients. We have shown that the RPE are critical for de novo serine synthesis in the mouse retina. To determine if serine dysregulation in RPE leads to disease in MacTel patients, we generated a library of iPSC-derived RPE from a cohort of MacTel patients and controls. We also generated CRISPR-modified cell lines where a PHGDH coding variant linked to MacTel was introduced into unaffected control lines. The same variant was corrected in MacTel patient lines. RPE were assayed for expression profiles, metabolic flux, and cellular function.

Methods : We generated and validated iPSCs from MacTel patients, unaffected family members, and unaffected controls. We used CRISPR/Cas9 to introduce PHGDH variants into control patient lines, and correct PHGDH variants in patient lines with inherent PHGDH variants. We probed for defects in serine metabolism using stable isotope tracing of carbon-13 labeled glucose. We further tested for metabolic dysfunction in bioenergetic assays using Seahorse to measure cellular respiration under various stressors.

Results : We show that a PHGDH variant found in MacTel patients is both necessary and sufficient to cause a significant reduction in serine biosynthesis and secretion in RPE cells. The level of serine reduction is also sufficient to induce accumulation of atypical lipids associated with neurodegeneration. A subset of patient RPE without a known causative mutation also show reduced serine biosynthesis when compared to family members. Seahorse assays show a serine-independent defect in mitochondrial respiration in MacTel patient RPE.

Conclusions : We show a possible cell-specific disease mechanism for MacTel by linking a newly discovered disease-linked genetic variant to reduced serine biosynthesis and lipid defects in patient RPE. We further demonstrate similar defects in MacTel patient RPE without the disease-linked PHGDH variant and suggest a serine-independent MacTel phenotype in mitochondrial function in the greater population.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.