Mitochondria must maintain adequate amounts of metabolites for protective and biosynthetic functions. However, how mitochondria sense the abundance of metabolites and regulate metabolic homeostasis is not well understood. In this work, we focused on glutathione (GSH), a critical redox metabolite in mitochondria, and identified a feedback mechanism that controls its abundance through the mitochondrial GSH transporter, SLC25A39. Under physiological conditions, SLC25A39 is rapidly degraded by mitochondrial protease AFG3L2. Depletion of GSH dissociates AFG3L2 from SLC25A39, causing a compensatory increase in mitochondrial GSH uptake. Genetic and proteomic analyses identified a putative iron-sulfur cluster in the matrix-facing loop of SLC25A39 as essential for this regulation, coupling mitochondrial iron homeostasis to GSH import. Altogether, our work revealed a paradigm for the autoregulatory control of metabolic homeostasis in organelles.

Measuring quality at varying levels of the healthcare system requires attribution, a process of determining the patients and services for which each level is responsible. However, it is important to ensure that attribution approaches are equitable; otherwise, individuals may be assigned differentially based upon social determinants of health. First, we used Medicaid claims (2010–2018) from Michigan to assess the proportion of children with sickle cell anemia that had less than 12 months enrollment within a single Medicaid health plan and could therefore not be attributed to a specific health plan. Second, we used the Medicaid Analytic eXtract (MAX) data (2008–2009) from 26 states to simulate adapting the 30-Day Pediatric All-Condition Readmission measure to the Accountable Care Organization (ACO) level and examined the proportion of readmissions that could not be attributed. For the sickle cell measure, an average of 300 children with sickle cell anemia were enrolled in Michigan Medicaid each year. The proportion of children that could not be attributed to a Medicaid health plan ranged from 12.2% to 89.0% across years. For the readmissions measure, of the 1,051,365 index admissions, 22% were excluded in the ACO-level analysis because of being unable to attribute the patient to a health plan for the 30 days post discharge. When applying attribution models, it is essential to consider the potential to induce health disparities. Differential attribution may have unintentional consequences that deepen health disparities, particularly when considering incentive programs for health plans to improve the quality of care.

Mitochondrial electron transport chain (ETC) dysfunction elevates the NADH/NAD⁺ ratio to cause metabolic derangements. Here we describe a protocol to measure the NADH/NAD⁺ ratio and analyze the rewiring of glucose metabolism using [4-²H]-glucose, [3-²H]-glucose, and [U-¹³C]-glucose in ETC-inhibited human cancer cells. We also describe a protocol to analyze the NADH/NAD⁺ ratio-sensitive metabolites in mouse plasma and mouse liver following phenformin treatment. These protocols comprehensively analyze the metabolic derangements resulting from increased NADH/NAD⁺ ratio in in vitro and in vivo models.