Investigating PHGDH deficiency inborn error metabolism in vitro and in vivo using gene regulation and metabolic tracing approaches

and Abstract Inborn errors of metabolism (IEMs) are a diverse group of over 1,000 congenital disorders caused by enzymatic mutation, the vast majority of which prominently affect the nervous system. Phosphoglycerate dehydrogenase (PHGDH) deficiency is a rare IEM caused by loss-of-function mutations in PHGDH, the rate- limiting enzyme in the de novo serine biosynthesis pathway. PHGDH deficiency presents with a clinically heterogeneous spectrum ranging from lethal neonatal Neu-Laxova Syndrome to nonlethal infantile-onset manifestations. While it is understood that PHGDH loss leads to serine and glycine depletion, the mechanisms linking enzyme dysfunction to nervous system phenotypes remain poorly defined. This project investigates how PHGDH loss disrupts metabolic homeostasis during nervous system development by disentangling its dual effects on serine-driven one-carbon, and central carbon metabolism. Preliminary data using [U-13C]glucose tracing in patient-derived PHGDH-deficient cells reveal not only reduced serine synthesis, but also increased pyruvate synthesis and TCA cycle turnover, potentially exacerbating oxidative stress through reactive oxygen species (ROS) overproduction. We hypothesize PHGDH deficiency not only inhibits serine/glycine synthesis, and also increases TCA cycle turnover, predisposing cells to oxidative damage while simultaneously driving global dysregulation of energy metabolism. In Aim 1, we will use inducible PHGDH expression systems in patient-derived fibroblast cell lines and [13C] tracers to define how varying PHGDH levels alter metabolic flux and ROS generation, and test interventions that buffer oxidative stress. In Aim 2, we will characterize the developmental consequences of PHGDH loss in newly developed humanized PHGDH deficiency mouse models. We will leverage spatial metabolic imaging and advanced microscopy to characterize the metabolic functions of PHGDH during nervous system development. Lastly, we will test exogenous, gestational serine supplementation strategies to assess tissue-specific metabolic vulnerability and embryonic lethality rescue in development. This work will provide fundamental insights into how PHGDH loss causes metabolic pathway disruptions leading to nervous system developmental defects. Our work will establish a framework for applying in vitro and in vivo targeted metabolic tracer and gene regulation approaches to study and potentially treat PHGDH deficiency and other IEMs. This proposal comprises of a rigorous and reproducible research strategy and comprehensive training plan that will prepare me to be an independent principal investigator at a tier 1 research institute. This research project will incorporate training in mass spectrometry, isotope tracer analysis, gene regulation, and molecular genetics. The University of Florida, in combination with the exceptional joint mentorship of Dr. Matthew Merritt and Dr. Eric Wang, provides an outstanding research environment that will enable my professional growth and completion of this proposed research. Project Number: 1F31HD121394-01 | Fiscal Year: 2025 | NIH Institute/Center: Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD) | Principal Investigator: Max Glanz | Institution: UNIVERSITY OF FLORIDA, GAINESVILLE, FL | Award Amount: $49,538 | Activity Code: F31 | Study Section: Special Emphasis Panel[ZRG1 F05-D (21)] View on NIH RePORTER: https://reporter.nih.gov/project-details/1F31HD12139401