Jeffrey C. Rathmell

The proliferation and development of activated B lymphocytes into antibody (Ab)-secreting plasma cells are critical determinants of humoral immunity and sustain certain autoimmune diseases. Specific pathways and substrate supplies in intermediary metabolism influence lymphocyte differentiation and function. We now show that although stringent restriction of glutamine supply decreases B cell proliferation and differentiation into plasma cells, glutaminolysis – a major metabolic use of this amino acid – was only conditionally crucial in B cells and the Ab responses derived from them. Gls , the gene encoding the main glutaminase of lymphocytes, promoted anti-NP Ab responses at the primary and recall phases if either glucose uptake into mouse B cells or pyruvate into their mitochondria was also impaired. Otherwise, it was dispensable. This synthetic auxotrophy, that is, the conditional requirement of glutaminase for processes in addition to survival, involved supporting progressive expansion of mitochondrial respiration followed by plasma cell differentiation. Surprisingly, impairment of glutaminase and the mitochondrial pyruvate channel decreased IL-21 stimulation of STAT3 phosphorylation as well as interferon stimulation of STAT1 activation. Together, our findings establish a powerful collaboration of metabolic pathways in programming increased respiration and the development of Ab-secreting cells, but also reveal modulation of cytokine receptor signaling by metabolism.

The efficiencies with which activated B lymphocytes proliferate and develop into antibody (Ab)-secreting plasma cells are critical determinants of adaptive humoral immunity and sustain certain autoimmune diseases. Specific pathways in intermediary metabolism, or their substrate supply, influence lymphocyte differentiation and function. We now show that although stringent restriction of glutamine supply decreases proliferation and differentiation of B cells into plasma cells, glutaminolysis - a major means of metabolism of this amino acid - was only conditionally crucial in B cells and the Ab responses derived from them. Strikingly, Gls, the gene encoding the main glutaminase of lymphocytes, promoted anti-NP Ab responses at the primary and recall phases if either glucose uptake into B cells or pyruvate into their mitochondria was also impaired but otherwise was dispensable. This synthetic auxotrophy, i.e., conditional requirement of glutaminase for processes in addition to survival and proliferation, involved support to a progressive expansion of mitochondrial respiration followed by plasma cell differentiation. Surprisingly, impairment of glutaminase and the mitochondrial pyruvate channel decreased IL-21 stimulation of STAT3 phosphorylation as well as interferon stimulation of STAT1 activation. Together, our findings establish not only a powerful collaboration of metabolic pathways in programming increased respiration and the development of Ab-secreting cells, but also reveal modulation of cytokine receptor signaling by metabolism.

The liver-α cell axis is a finely tuned biological rheostat that regulates whole body amino acid availability. Pancreatic α cells secrete glucagon that regulates amino acid catabolism through gluconeogenesis and ureagenesis, yet the mechanisms linking amino acid levels to α cell growth and function are not fully understood. Here, we identify glutaminase, the enzyme that catalyzes glutamine catabolism, as a critical α cell regulator. Glutaminase is highly enriched in α cells across species. α cell expression of glutaminase is required for nutrient-dependent mTORC1 activation, suppression of AMPK signaling, and sustained expression of the glutamine transporter SLC38A5. This establishes a feed-forward loop linking glutamine metabolism to amino acid sensing and growth. Reduced glutaminase activity impairs dynamic glucagon secretion in response to low glucose and amino acids. Together, these findings highlight the importance of glutamine metabolism in α cell growth and hormone secretion and suggest it may play a role in α cell adaptation to hyperaminoacidemia.

The imbalance of pro-inflammatory and immunosuppressive constituents in the tumor microenvironment (TME) significantly dictates cancer progression, immune invasion, and treatment response. Immunosuppressive regulatory T cells (Tregs) are associated with decreased disease-free survival due to their ability to suppress anti-tumor immunity. Notably, in response to environmental cues, Tregs display functional and phenotypic plasticity with inflammatory helper T cell subsets. Given their predominant role in sustaining immune suppression, it is remarkable that multiple developmental pathways converge to regulate Treg development and function. In this study, we designed, validated, and employed a novel genetically modified mouse model to conditionally ablate the Hedgehog (Hh) gene, Gli2, specifically in Tregs. Ablation of Gli2 activity in Tregs significantly reduced tumor burden, impaired Treg suppressive function, and shifted the transcriptional balance of Foxo3 and Rorγt, transcription factors essential for Tregs and Th17 cells. Spatial mapping highlighted that Gli2 ablation in Tregs enhances the immunogenicity of the tumor and promotes a pro-inflammatory milieu of the TME. This was underscored by a higher tumor immune signature score and enhanced infiltration of cytotoxic CD8 ⁺ T cells into the tumor. These findings highlight Hh/Gli2 signaling in Tregs as a mechanistic regulator of immunogenicity in the TME and a potential therapeutic target to prime tumors for enhanced responsiveness for adjuvant treatments.

Immune cell populations in tumors and tumor-adjacent mammary adipose tissue (mAT) analyzed by single-cell RNA sequencing.

Mammary adipose tissue lipid associated macrophage marker gene expression and correlation with tumor mass.

Individual tumor volumes over time and oAT and mAT mass at study endpoint.

IgG and anti-PD-1 treatment in lean, obese, and weight loss mice.

Food intake and pre-diet switch glucose tolerance in lean, obese, and weight loss mice.

Checkpoint inhibitors have transformed cancer treatment, yet predicting responses remains challenging. Mitochondrial quality decreases in tumor infiltrating lymphocytes and correlates with impaired antitumor immunity in animal models. Mitochondrial membrane potential (MMP) increases with T cell activation and may also indicate cellular dysfunction. Here, we assessed the MMP of tumor-associated T cells as an indicator of cell phenotypes and immunotherapy responses in non-small cell lung carcinoma and clear cell renal cell carcinoma patients. Primary tumors were collected followed by analysis of peripheral blood mononuclear cells prior to and after 3 wk on treatment with immune checkpoint inhibitors (ICIs). Peripheral blood mononuclear T cells were analyzed for MMP using tetramethylrhodamine ethyl ester (TMRE) and sorted into high and low populations. TCRβ and single-cell RNA sequencing of primary tumors identified and characterized peripheral blood T cell clones associated with the tumor microenvironment. As anticipated, ICI therapy increased the frequency of effector T cells in patients who experienced clinical benefit. TMREhigh peripheral blood T cells with tumor-matching TCRβ sequences had elevated oxidative phosphorylation gene signatures. Gene signatures of stress and exhaustion, such as Tigit and Cmc1, were also elevated in the TMREhigh CD8 T cell populations, while gene expression patterns in TMRElow cells suggested mitochondrial fitness and cell longevity. Importantly, clinical benefit from ICIs was negatively correlated with the TMREhigh CD8 T cell gene expression signature. These findings highlight a T cell population characterized by elevated MMP that correlates with exhaustion-like transcriptional states and poor response to immunotherapy.