Dissecting the Glycolytic Requirements of Human T Cells via G6PC3 Deficiency

Matthew Stier – Vanderbilt University Medical Center, Nashville, TN, USA; Christine Mariskanish – Vanderbilt University Medical Center, Nashville, TN, USA; Jerab Perez Caraballo – Vanderbilt University Medical Center, Nashville, TN, USA; Megan Dobrose – Vanderbilt University Medical Center, Nashville, TN, USA; Edgar Alejandro Medina-Torres – National Institute of Pediatrics, Health secretariat, Mexico City, Mexico; Selma Cecilia Scheffler Mendoza – National Institute of Pediatrics, Health secretariat, Mexico City, Mexico; James Connelly – Vanderbilt University Medical Center, Nashville, TN, USA; Luis Ignacio Gonzalez-Granado – University Hospital 12 de Octubre, Madrid, Spain; Diana Andrea Herrera Sánchez – National Medical Center XXI Century, Mexico City, Mexico; Patricia O’Farrill Romanillos – National Medical Center XXI Century, Mexico City, Mexico; Kelly Walkovich – University of Michigan, Ann Arbor, MI, USA; Jeffrey Rathmell – Vanderbilt University Medical Center, Nashville, TN, USA; Saul Oswaldo Lugo-Reyes – National Institute of Pediatrics, Health Secretariat, Mexico City, Mexico; Ruben Martinez-Barricarte – Vanderbilt University Medical Center, Nashville, TN, USA

Abstract Text: The glycolytic requirements of T cells in vivo in humans remain elusive, in part, due to the impossibility of prolonged inhibition of glycolysis. We tackle this limitation by studying glucose-6-phosphatase catalytic subunit 3 (G6PC3) deficiency, a monogenic immunometabolic disorder that causes intracellular accumulation of the metabolite 1,5-anhydroglucitol-6-phosphate (1,5-AG6P) in immune cells.
We demonstrate that 1,5-AG6P accumulation markedly impairs glycolytic activities in G6PC3 CRISPR/Cas9 knockout Jurkat T cells and in primary T cells from G6PC3-deficient patients. These findings and our unique cohort of patients with G6PC3 deficiency allowed us to examine how impaired glycolysis impact human T cell responses. Multidimensional analysis using CyTOF shows that G6PC3-deficient patients display lower frequencies of naïve T cells with a concomitant increase in the effector memory T cell and TEMRA proportions in both CD4+ and CD8+ compartments. Patients also have an increased frequency of TH2/TH17 (CD4+CD45RA-CXCR3-) cells and indications of impaired thymic function, showcasing the importance of glycolysis for T cell development and differentiation. Furthermore, we show the correlation between T cell glycolytic activities and PD-1 expression, since patient memory T cells exhibit elevated PD-1 expression, and CD3/CD28 stimulation induces a significantly greater upregulation of PD-1 expression on CD4+ and CD8+ T cells from patients than those from healthy donors. An integrative analysis of scRNA-seq and scTCR-seq data shed light on potential mechanisms underlying these observations.
Collectively, this study highlights the importance of studying rare genetic diseases as an approach to interrogate the links between cellular metabolism and human immune cell fate and function.