The role of tristetraprolin in inflammation and disease

Untersuchungen über die Kontrolle der Genexpression durch transkriptionelle und posttranskriptionelle Mechanismen sind wichtig für das weiterführende Verständnis humaner Pathologien. Während die transkriptionelle Ebene in der Vergangenheit verstärkt erforscht wurde, wenden viele Arbeitsgruppen in den letzten Jahren ihre Aufmerksamkeit posttranskriptionellen Mechanismen zu, die mRNA-Reifung, -Transport, -Stabilität und -Translation steuern. Tristetraprolin (TTP) bindet AU-reiche Sequenzen im 3’UTR vieler mRNAs und leitet damit den raschen Abbau dieser RNAs ein. Tiermodelle sowie in vitro Studien haben gezeigt, dass TTP eine wichtige Kontrollfunktion im Immunsystem hat. Mäuse deren TTP-Gen entfernt wurde entwickeln daher verschiedene Formen chronischer Entzündung wie Arthritis. Diese Entzündungsreaktionen werden auf die erhöhte Stabilität der TNFα mRNA zurückgeführt. Da TNFα die Gesundheit dieser Mäuse stark beeinflusst, konnten bisher keine weiterführenden Studien zur Rolle von TTP in anderen Krankheiten durchgeführt werden. Wir entwickelten daher eine Maus, deren TTP-Gen gewebsspezifisch ausgeschaltet werden kann. Dadurch konnten wir zeigen, dass Tiere die kein myeloides TTP produzieren, anfälliger für LPS-induzierten endotoxischen Schock sind. Um die Funktion von TTP besser zu verstehen, versuchten wir weitere Gene zu identifizieren, deren mRNA-Stabilität durch TTP gesteuert wird. Im Zuge dessen konnten wir zeigen, dass TTP Teil einer Feedbackschleife während entzündlicher Reaktionen ist. In dieser Schleife wird die Entzündung als auch die Expression von TTP durch die p38 MAP-Kinase initiiert. Gleichzeitig phosphoryliert p38 MAP-Kinase das TTP Protein wodurch es inaktiv wird und viele mRNAs stabilisiert werden. Im späteren Verlauf der Entzündung wird p38 MAP-Kinase allmählich durch Interleukin-10 (IL-10) und die durch IL-10 verstärkt produzierte p38 MAP-Kinase Phosphatase DUSP1 blockiert. Nun kommt es zur TTP Aktivierung und zum Abbau eines großen Anteils an LPS-induzierten mRNAs.Uncovering control of gene expression by transcriptional and posttranscriptional mechanisms is of uppermost importance for the understanding of human pathologies. While the transcriptional level has been extensively studied in the past, in recent years more and more studies focus on the control of RNA maturation, transport, stability and translation. Among others, tristetraprolin (TTP) has been described to bind to AU-rich elements (AREs) within the 3’UTR of several mRNAs mediating rapid degradation. Animal models as well as in vitro studies revealed a profound role of TTP in controlling immune homeostasis by regulating mRNA stability of inflammatory mediators. Conventional TTP knockout mice therefore develop various pathologies like rheumatoid arthritis resulting from increased TNFα mRNA stability. However the function of TTP in different states of diseases could not be addressed properly in these animals as they suffer from chronic inflammation caused by dominant TNFα overproduction. We therefore created a conditional TTP knockout mouse-line in order to analyze the role of TTP in acute inflammation. Animals deleted for TTP in myeloid cells appeared to be more susceptible to LPS driven endotoxic shock, pointing out the importance of TTP in limiting the inflammatory response. So far only a limited number of TTP-targets has been described. In order to explain the phenotype seen in different disease models we tried to identify new TTP-targets and how TTP is regulated during inflammation. We and others could show that the main stress-kinase p38 MAPK induces TTP expression but also phosphorylates the TTP protein therefore blocking its function. Upcoming Interleukin-10 (IL-10) signaling at later stages of inflammation activates the phosphates DUSP1. DUSP1 in turn inhibits TTP phosphorylation by p38 MAPK thereby engaging TTP activity. TTP therefore leads to the removal of a large proportion of unstable mRNAs in a p38 MAPK dependent way as part of a global acting negative feedback loop

Type I Interferon Production Induced by Streptococcus pyogenes-Derived Nucleic Acids Is Required for Host Protection

Streptococcus pyogenes is a Gram-positive human pathogen that is recognized by yet unknown pattern recognition receptors (PRRs). Engagement of these receptor molecules during infection with S. pyogenes, a largely extracellular bacterium with limited capacity for intracellular survival, causes innate immune cells to produce inflammatory mediators such as TNF, but also type I interferon (IFN). Here we show that signaling elicited by type I IFNs is required for successful defense of mice against lethal subcutaneous cellulitis caused by S. pyogenes. Type I IFN signaling was accompanied with reduced neutrophil recruitment to the site of infection. Mechanistic analysis revealed that macrophages and conventional dendritic cells (cDCs) employ different signaling pathways leading to IFN-beta production. Macrophages required IRF3, STING, TBK1 and partially MyD88, whereas in cDCs the IFN-beta production was fully dependent on IRF5 and MyD88. Furthermore, IFN-beta production by macrophages was dependent on the endosomal delivery of streptococcal DNA, while in cDCs streptococcal RNA was identified as the IFN-beta inducer. Despite a role of MyD88 in both cell types, the known IFN-inducing TLRs were individually not required for generation of the IFN-beta response. These results demonstrate that the innate immune system employs several strategies to efficiently recognize S. pyogenes, a pathogenic bacterium that succeeded in avoiding recognition by the standard arsenal of TLRs

T Cell Cancer Therapy Requires CD40-CD40L Activation of Tumor Necrosis Factor and Inducible Nitric-Oxide-Synthase-Producing Dendritic Cells

Effective cancer immunotherapy requires overcoming immunosuppressive tumor microenvironments. We\ua0found that local nitric oxide (NO) production by tumor-infiltrating myeloid cells is important for adoptively transferred CD8(+) cytotoxic T\ua0cells to destroy tumors. These myeloid cells are phenotypically similar to inducible nitric oxide synthase (NOS2)- and tumor necrosis factor (TNF)-producing dendritic cells (DC), or Tip-DCs. Depletion of immunosuppressive, colony stimulating factor 1 receptor (CSF-1R)-dependent arginase 1(+) myeloid cells enhanced NO-dependent tumor killing. Tumor elimination via NOS2 required the CD40-CD40L pathway. We also uncovered a strong correlation between survival of colorectal cancer patients and NOS2, CD40, and TNF expression in their tumors. Our results identify a network of pro-tumor factors that can be targeted to boost cancer immunotherapies

The RNA-binding protein tristetraprolin schedules apoptosis of pathogen-engaged neutrophils during bacterial infection

Protective responses against pathogens require a rapid mobilization of resting neutrophils and the timely removal of activated ones. Neutrophils are exceptionally short-lived leukocytes, yet it remains unclear whether the lifespan of pathogen-engaged neutrophils is regulated differently from that in the circulating steady-state pool. Here, we have found that under homeostatic conditions, the mRNA-destabilizing protein tristetraprolin (TTP) regulates apoptosis and the numbers of activated infiltrating murine neutrophils but not neutrophil cellularity. Activated TTP-deficient neutrophils exhibited decreased apoptosis and enhanced accumulation at the infection site. In the context of myeloid-specific deletion of Ttp, the potentiation of neutrophil deployment protected mice against lethal soft tissue infection with Streptococcus pyogenes and prevented bacterial dissemination. Neutrophil transcriptome analysis revealed that decreased apoptosis of TTP-deficient neutrophils was specifically associated with elevated expression of myeloid cell leukemia 1 (Mcl1) but not other antiapoptotic B cell leukemia/ lymphoma 2 (Bcl2) family members. Higher Mcl1 expression resulted from stabilization of Mcl1 mRNA in the absence of TTP. The low apoptosis rate of infiltrating TTP-deficient neutrophils was comparable to that of transgenic Mcl1-overexpressing neutrophils. Our study demonstrates that posttranscriptional gene regulation by TTP schedules the termination of the antimicrobial engagement of neutrophils. The balancing role of TTP comes at the cost of an increased risk of bacterial infections

TNF Counterbalances the Emergence of M2 Tumor Macrophages

Cancer can involve non-resolving, persistent inflammation where varying numbers of tumor-associated macrophages (TAMs) infiltrate and adopt different activation states between anti-tumor M1 and pro-tumor M2 phenotypes. Here, we resolve a cascade causing differential macrophage phenotypes in the tumor microenvironment. Reduction in TNF mRNA production or loss of type I TNF receptor signaling resulted in a striking pattern of enhanced M2 mRNA expression. M2 gene expression was driven in part by IL-13 from eosinophils co-recruited with inflammatory monocytes, a pathway that was suppressed by TNF. Our data define regulatory nodes within the tumor microenvironment that balance M1 and M2 populations. Our results show macrophage polarization in cancer is dynamic and dependent on the balance between TNF and IL-13, thus providing a strategy for manipulating TAMs

Tristetraprolin is required for full anti-inflammatory response of murine macrophages to IL-10.

IL-10 is essential for inhibiting chronic and acute inflammation by decreasing the amounts of proinflammatory cytokines made by activated macrophages. IL-10 controls proinflammatory cytokine and chemokine production indirectly via the transcription factor Stat3. One of the most physiologically significant IL-10 targets is TNF-alpha, a potent proinflammatory mediator that is the target for multiple anti-TNF-alpha clinical strategies in Crohn's disease and rheumatoid arthritis. The anti-inflammatory effects of IL-10 seem to be mediated by several incompletely understood transcriptional and posttranscriptional mechanisms. In this study, we show that in LPS-activated bone marrow-derived murine macrophages, IL-10 reduces the mRNA and protein levels of TNF-alpha and IL-1alpha in part through the RNA destabilizing factor tristetraprolin (TTP). TTP is known for its central role in destabilizing mRNA molecules containing class II AU-rich elements in 3' untranslated regions. We found that IL-10 initiates a Stat3-dependent increase of TTP expression accompanied by a delayed decrease of p38 MAPK activity. The reduction of p38 MAPK activity releases TTP from the p38 MAPK-mediated inhibition, thereby resulting in diminished mRNA and protein levels of proinflammatory cytokines. These findings establish that TTP is required for full responses of bone marrow-derived murine macrophages to IL-10