CARMIL2 deficiency disrupts activation-induced metabolic reprogramming in T cells and is partially rescued by glutamine supplementation

Background: T-cell activation requires signaling through the T-cell receptor and costimulatory molecules, including CD28, triggering metabolic reprogramming to support growth and proliferation of the activating T cell. CARMIL2, a scaffold protein, facilitates CD28-mediated signaling. Individuals with CARMIL2 mutations experience inborn errors of immunity, leading to T-cell dysfunction and severe infectious and inflammatory comorbidities. However, how CARMIL2 deficiency impacts T-cell metabolic reprogramming remains unknown. Objective: We sought to investigate how CARMIL2 deficiency affects activation-induced metabolic reprogramming in T cells. Methods: CD4⁺ T cells were isolated from patients with CARMIL2 deficiency and matched healthy controls. A transcriptomic profile was analyzed by bulk RNA sequencing and whole-cell metabolomics by LC-MS/MS. Activation markers and signaling pathways were measured by flow cytometry. These approaches informed identification of specific amino acids for rescue experiments. Results: Nine patients with CARMIL2 deficiency and 16 age- and sex-matched healthy controls were recruited. RNA sequencing of CD4⁺ T cells revealed decreased expression of genes associated with metabolic activity, including mTOR signaling, glycolysis, 1-carbon metabolism, and glutamine metabolism. Whole-cell metabolomics reinforced these results and highlighted glutamine deficiency as a potential driver of the observed metabolic phenotype. Glutamine supplementation restored NF-κB and mTOR activity, as measured by p-65 and RPS6 phosphorylation, respectively, and upregulated the expression of IL17A in CARMIL2-mutated CD4⁺ T cells. Conclusion: CARMIL2 deficiency disrupts T-cell metabolic reprogramming and was partially rescued ex vivo with glutamine supplementation. These findings highlight a potential therapeutic approach targeting metabolism to improve immune function in individuals with CARMIL2 deficiency.