SUMOylation controls Hu antigen R posttranscriptional activity in liver cancer.

The posttranslational modification of proteins critically influences many biological processes and is a key mechanism that regulates the function of the RNA-binding protein Hu antigen R (HuR), a hub in liver cancer. Here, we show that HuR is SUMOylated in the tumor sections of patients with hepatocellular carcinoma in contrast to the surrounding tissue, as well as in human cell line and mouse models of the disease. SUMOylation of HuR promotes major cancer hallmarks, namely proliferation and invasion, whereas the absence of HuR SUMOylation results in a senescent phenotype with dysfunctional mitochondria and endoplasmic reticulum. Mechanistically, SUMOylation induces a structural rearrangement of the RNA recognition motifs that modulates HuR binding affinity to its target RNAs, further modifying the transcriptomic profile toward hepatic tumor progression. Overall, SUMOylation constitutes a mechanism of HuR regulation that could be potentially exploited as a therapeutic strategy for liver cancer.

Lactate limits CNS autoimmunity by stabilizing HIF-1α in dendritic cells.

Dendritic cells (DCs) have a role in the development and activation of self-reactive pathogenic T cells1,2. Genetic variants that are associated with the function of DCs have been linked to autoimmune disorders3,4, and DCs are therefore attractive therapeutic targets for such diseases. However, developing DC-targeted therapies for autoimmunity requires identification of the mechanisms that regulate DC function. Here, using single-cell and bulk transcriptional and metabolic analyses in combination with cell-specific gene perturbation studies, we identify a regulatory loop of negative feedback that operates in DCs to limit immunopathology. Specifically, we find that lactate, produced by activated DCs and other immune cells, boosts the expression of NDUFA4L2 through a mechanism mediated by hypoxia-inducible factor 1α (HIF-1α). NDUFA4L2 limits the production of mitochondrial reactive oxygen species that activate XBP1-driven transcriptional modules in DCs that are involved in the control of pathogenic autoimmune T cells. We also engineer a probiotic that produces lactate and suppresses T cell autoimmunity through the activation of HIF-1α-NDUFA4L2 signalling in DCs. In summary, we identify an immunometabolic pathway that regulates DC function, and develop a synthetic probiotic for its therapeutic activation.

Engineered probiotics limit CNS autoimmunity by stabilizing HIF-1α in dendritic cells.

Dendritic cells (DCs) control the generation of self-reactive pathogenic T cells. Thus, DCs are considered attractive therapeutic targets for autoimmune diseases. Using single-cell and bulk transcriptional and metabolic analyses in combination with cell-specific gene perturbation studies we identified a negative feedback regulatory pathway that operates in DCs to limit immunopathology. Specifically, we found that lactate, produced by activated DCs and other immune cells, boosts NDUFA4L2 expression through a mechanism mediated by HIF-1α. NDUFA4L2 limits the production of mitochondrial reactive oxygen species that activate XBP1-driven transcriptional modules in DCs involved in the control of pathogenic autoimmune T cells. Moreover, we engineered a probiotic that produces lactate and suppresses T-cell autoimmunity in the central nervous system via the activation of HIF-1α/NDUFA4L2 signaling in DCs. In summary, we identified an immunometabolic pathway that regulates DC function, and developed a synthetic probiotic for its therapeutic activation.

From Nucleus to Organs: Insights of Aryl Hydrocarbon Receptor Molecular Mechanisms.

The aryl hydrocarbon receptor (AHR) is a markedly established regulator of a plethora of cellular and molecular processes. Its initial role in the detoxification of xenobiotic compounds has been partially overshadowed by its involvement in homeostatic and organ physiology processes. In fact, the discovery of its ability to bind specific target regulatory sequences has allowed for the understanding of how AHR modulates such processes. Thereby, AHR presents functions in transcriptional regulation, chromatin architecture modifications and participation in different key signaling pathways. Interestingly, such fields of influence end up affecting organ and tissue homeostasis, including regenerative response both to endogenous and exogenous stimuli. Therefore, from classical spheres such as canonical transcriptional regulation in embryonic development, cell migration, differentiation or tumor progression to modern approaches in epigenetics, senescence, immune system or microbiome, this review covers all aspects derived from the balance between regulation/deregulation of AHR and its physio-pathological consequences.

Identification of environmental factors that promote intestinal inflammation.

Genome-wide association studies have identified risk loci linked to inflammatory bowel disease (IBD)1-a complex chronic inflammatory disorder of the gastrointestinal tract. The increasing prevalence of IBD in industrialized countries and the augmented disease risk observed in migrants who move into areas of higher disease prevalence suggest that environmental factors are also important determinants of IBD susceptibility and severity2. However, the identification of environmental factors relevant to IBD and the mechanisms by which they influence disease has been hampered by the lack of platforms for their systematic investigation. Here we describe an integrated systems approach, combining publicly available databases, zebrafish chemical screens, machine learning and mouse preclinical models to identify environmental factors that control intestinal inflammation. This approach established that the herbicide propyzamide increases inflammation in the small and large intestine. Moreover, we show that an AHR-NF-κB-C/EBPß signalling axis operates in T cells and dendritic cells to promote intestinal inflammation, and is targeted by propyzamide. In conclusion, we developed a pipeline for the identification of environmental factors and mechanisms of pathogenesis in IBD and, potentially, other inflammatory diseases.

Liver regeneration after partial hepatectomy is improved in the absence of aryl hydrocarbon receptor.

The liver is among the few organs having the ability to self-regenerate in response to a severe damage compromising its functionality. The Aryl hydrocarbon receptor (Ahr) is a transcription factor relevant for the detoxification of xenobiotics but also largely important for liver development and homeostasis. Hence, liver cell differentiation is developmentally modulated by Ahr through the controlled expression of pluripotency and stemness-inducing genes. Here, 2/3 partial hepatectomy (PH) was used as a clinically relevant approach to induce liver regeneration in Ahr-expressing (Ahr+/+) and Ahr-null (Ahr-/-) mice. Ahr expression and activity were early induced after 2/3 PH to be gradually downmodulated latter during regeneration. Ahr-/- mice triggered liver regeneration much faster than AhR+/+ animals, although both reached full regeneration at the latest times. At initial stages after PHx, earlier regenerating Ahr-/- livers had upregulation of cell proliferation markers and increased activation of signalling pathways related to stemness such as Hippo-YAP and Wnt/ß-catenin, concomitantly with the induction of pro-inflammatory cytokines TNFa, IL6 and p65. These phenotypes, together with the improved metabolic adaptation of Ahr-/- mice after PHx and their induced sustained cell proliferation, could likely result from the expansion of undifferentiated stem cells residing in the liver expressing OCT4, SOX2, KLF4 and NANOG. We propose that Ahr needs to be induced early during regeneration to fine-tune liver regrowth to physiological values. Since Ahr deficiency did not result in liver overgrowth, its transient pharmacological inhibition could serve to improve liver regeneration in hepatectomized and transplanted patients and in those exposed to damaging liver toxins and carcinogens.

Aryl hydrocarbon receptor blocks aging-induced senescence in the liver and fibroblast cells.

Aging impairs organismal homeostasis leading to multiple pathologies. Yet, the mechanisms and molecular intermediates involved are largely unknown. Here, we report that aged aryl hydrocarbon receptor-null mice (AhR-/-) had exacerbated cellular senescence and more liver progenitor cells. Senescence-associated markers ß-galactosidase (SA-ß-Gal), p16Ink4a and p21Cip1 and genes encoding senescence-associated secretory phenotype (SASP) factors TNF and IL1 were overexpressed in aged AhR-/- livers. Chromatin immunoprecipitation showed that AhR binding to those gene promoters repressed their expression, thus adjusting physiological levels in AhR+/+ livers. MCP-2, MMP12 and FGF secreted by senescent cells were overproduced in aged AhR-null livers. Supporting the relationship between senescence and stemness, liver progenitor cells were overrepresented in AhR-/- mice, probably contributing to increased hepatocarcinoma burden. These AhR roles are not liver-specific since adult and embryonic AhR-null fibroblasts underwent senescence in culture, overexpressing SA-ß-Gal, p16Ink4a and p21Cip1. Notably, depletion of senescent cells with the senolytic agent navitoclax restored expression of senescent markers in AhR-/- fibroblasts, whereas senescence induction by palbociclib induced an AhR-null-like phenotype in AhR+/+ fibroblasts. AhR levels were downregulated by senescence in mouse lungs but restored upon depletion of p16Ink4a-expressing senescent cells. Thus, AhR restricts age-induced senescence associated to a differentiated phenotype eventually inducing resistance to liver tumorigenesis.

The aryl hydrocarbon receptor promotes differentiation during mouse preimplantational embryo development.

Mammalian embryogenesis is a complex process controlled by transcription factors that regulate the balance between pluripotency and differentiation. Transcription factor aryl hydrocarbon receptor (AhR) regulates OCT4/POU5F1 and NANOG, both essential controllers of pluripotency, stemness and early embryo development. Molecular mechanisms controlling OCT4/POU5F1 and NANOG during embryogenesis remain unidentified. We show that AhR regulates pluripotency factors and maintains the metabolic activity required for proper embryo differentiation. AhR-lacking embryos (AhR-/-) showed a pluripotent phenotype characterized by a delayed expression of trophectoderm differentiation markers. Accordingly, central pluripotency factors OCT4/POU5F1 and NANOG were overexpressed in AhR-/- embryos at initial developmental stages. An altered intracellular localization of these factors was observed in the absence of AhR and, importantly, Oct4 had an opposite expression pattern with respect to AhR from the two-cell stage to blastocyst, suggesting a negative regulation of OCT4/POU5F by AhR. We propose that AhR is a regulator of pluripotency and differentiation in early mouse embryogenesis.

Loss of Aryl Hydrocarbon Receptor Favors K-RasG12D-Driven Non-Small Cell Lung Cancer.

Non-small cell lung adenocarcinoma (NSCLC) bearing K-RasG12D mutations is one of the most prevalent types of lung cancer worldwide. Aryl hydrocarbon receptor (AHR) expression varies in human lung tumors and has been associated with either increased or reduced lung metastasis. In the mouse, Ahr also adjusts lung regeneration upon injury by limiting the expansion of resident stem cells. Here, we show that the loss of Ahr enhances K-RasG12D-driven NSCLC in mice through the amplification of stem cell subpopulations. Consistent with this, we show that K-RasG12D;Ahr-/- lungs contain larger numbers of cells expressing markers for both progenitor Clara (SCGB1A1 and CC10) and alveolar type-II (SFTPC) cells when compared to K-RasG12D;Ahr+/+-driven tumors. They also have elevated numbers of cells positive for pluripotent stem cells markers such as SOX2, ALDH1, EPCAM, LGR5 and PORCN. Typical pluripotency genes Nanog, Sox2 and c-Myc were also upregulated in K-RasG12D;Ahr-/- lung tumors as found by RNAseq analysis. In line with this, purified K-RasG12D/+;Ahr-/- lung cells generate larger numbers of organoids in culture that can subsequently differentiate into bronchioalveolar structures enriched in both pluripotency and stemness genes. Collectively, these data indicate that Ahr antagonizes K-RasG12D-driven NSCLC by restricting the number of cancer-initiating stem cells. They also suggest that Ahr expression might represent a good prognostic marker to determine the progression of K-RasG12D-positive NSCLC patients.