Biology subjects

Longworth, J.

Publications and source records attributed to Longworth, J..

3 recordsLinked to original sources

bioRxiv2023

A Th17 cell-intrinsic glutathione/mitochondrial-IL-22 axis protects against intestinal inflammation

Although the intestinal tract is a major site of reactive oxygen species (ROS) generation, the mechanisms by which antioxidant defense in gut T cells contribute to intestinal homeostasis are currently unknown. Here we show, using T cell-specific ablation of the catalytic subunit of glutamate cysteine ligase (Gclc), that the ensuing loss of glutathione (GSH) impairs the production of gut-protective IL-22 by Th17 cells within the lamina propria. Although Gclc ablation does not affect T cell cytokine secretion in the gut of mice at steady-state, infection with C. rodentium increases ROS, inhibits mitochondrial gene expression and mitochondrial function in Gclc-deficient Th17 cells. These mitochondrial deficits affect the PI3K/AKT/mTOR pathway, leading to reduced phosphorylation of the translation repressor 4E-BP1. As a consequence, the initiation of translation is restricted, resulting in decreased protein synthesis of IL-22. Loss of IL-22 results in poor bacterial clearance, enhanced intestinal damage, and high mortality. ROS-scavenging, reconstitution of IL-22 expression or IL-22 supplementation in vivo prevent the appearance of these pathologies. Our results demonstrate the existence of a previously unappreciated role for Th17 cell-intrinsic GSH coupling to promote mitochondrial function, IL-22 translation and signaling. These data reveal an axis that is essential for maintaining the integrity of the intestinal barrier and protecting it from damage caused by gastrointestinal infection. Executive summary- GSH-regulated Th17 cell-derived IL-22, but not IL-17 is required to maintain intestinal barrier integrity and to revent lethality following C. rodentium infection. - GCLC expression in IBD patients correlates positively with expression of genes related to gut integrity. - Gclc-deficient Th17 cells accumulate mitochondrial ROS, which is linked to impaired mitochondrial function, ysregulated PI3K/AKT/mTOR signaling and impaired translation of IL-22. - ROS-scavenging, IL-22 reconstitution or T cell-specific expression of IL-22 in Gclc-deficient T cells rescues utant mice from the lethal infection outcome in vivo.

immunology↗

bioRxiv2023

A multi-omics integrative approach unravels novel genes and pathways associated with senescence escape after targeted therapy in NRAS mutant melanoma

Therapy Induced Senescence (TIS) leads to sustained growth arrest of cancer cells. The associated cytostasis has been shown to be reversible and cells escaping senescence further enhance the aggressiveness of cancers. Together with targeted therapeutics, senolytics, specifically targeting senescent cancer cells, constitute a promising avenue for improved cancer treatments. Understanding how cancer cells evade senescence is needed to optimise the clinical benefits of this therapeutic approach. Here we characterised the response of three different NRAS mutant melanoma cell lines to a combination of CDK4/6 and MEK inhibitors over 33 days. Transcriptomic data show that all cell lines trigger a senescence programme coupled with strong induction of interferons. Kinome profiling revealed the activation of Receptor Tyrosine Kinases (RTKs) and enriched downstream signaling of neurotrophin, ErbB and insulin pathways. Characterisation of the miRNA interactome associates miR-211-5p with resistant phenotypes. Finally, iCELL-based integration of bulk and single-cell RNA-seq data identified biological processes perturbed during senescence, and predicts new genes involved in its escape. Overall, our data associate insulin signaling with persistence of a senescent phenotype and suggest a new role for interferon gamma in senescence escape through the induction of EMT and the activation of ERK5 signaling.

cancer biology↗

bioRxiv2020

Rub1/NEDD8, a ubiquitin-like modifier, is also a ubiquitin modifier

Of all ubiquitin-like small protein modifiers, Rub1/NEDD8 is the closest kin of ubiquitin in sequence and in structure. Despite their profound similarities, prevalence of ubiquitin and of Rub1 is starkly different: targets of ubiquitin modification reach into the thousands, whereas unique targets of Rub1/NEDD8 appear limited to one family of proteins, Cullins. This distinction is likely due to dedicated E1 activating enzymes that select either one or the other and relay the modifier until it is covalently attached to a target. To convert typical neddylation targets for modification by ubiquitin, and vice versa, we designed reciprocal substitutions at position 72 of Rub1 and of ubiquitin to render them substrates for activation by their non-cognate E1 activating enzymes. We found that this single amino acid is sufficient to distinguish between Ub and Rub1 in living cells, and determine their targets. Thus, modification of Cullins by UbR72T could compensate for loss of Rub1, even as it maintained its ability to polymerize and direct conjugates for degradation. Conversely, Rub1T72R activated by ubiquitin-activating enzyme entered into the ubiquitination cascade, however was not efficiently polymerized, essentially capping polyubiquitin chains. Upon shortage of free ubiquitin under stress, even native Rub1 spilled-over into the ubiquitinome suppressing polyubiquitination. By contrast, the need to maintain monomeric modifications on unique targets is a likely explanation for why the Rub1-activating enzyme strictly discriminates against ubiquitin. Swapping Rub1 and ubiquitin signals uncovered a reason for maintaining two separate pathways across eukaryotic kingdom.

biochemistry↗