T cell differentiation into distinct functional effector and inhibitory subsets is regulated, in part, by the cytokine environment present at the time of antigen recognition. Here, we show that ...hypoxia-inducible factor 1 (HIF-1), a key metabolic sensor, regulates the balance between regulatory T cell (Tᵣₑg) and TH17 differentiation. HIF-1 enhances TH17 development through direct transcriptional activation of RORγt and via tertiary complex formation with RORγt and p300 recruitment to the IL-17 promoter, thereby regulating TH17 signature genes. Concurrently, HIF-1 attenuates Tᵣₑg development by binding Foxp3 and targeting it for proteasomal degradation. Importantly, this regulation occurs under both normoxic and hypoxic conditions. Mice with HIF-1α-deficient T cells are resistant to induction of TH17-dependent experimental autoimmune encephalitis associated with diminished TH17 and increased Tᵣₑg cells. These findings highlight the importance of metabolic cues in T cell fate determination and suggest that metabolic modulation could ameliorate certain T cell-based immune pathologies.
Wastewater surveillance (WS), when coupled with advanced molecular techniques, offers near real-time monitoring of community-wide transmission of SARS-CoV-2 and allows assessing and mitigating ...COVID-19 outbreaks, by evaluating the total microbial assemblage in a community. Composite wastewater samples (24 h) were collected weekly from a manhole between December 2020 and November 2021 in Maryland, USA. RT-qPCR results showed concentrations of SARS-CoV-2 RNA recovered from wastewater samples reflected incidence of COVID-19 cases. When a drastic increase in COVID-19 was detected in February 2021, samples were selected for microbiome analysis (DNA metagenomics, RNA metatranscriptomics, and targeted SARS-CoV-2 sequencing). Targeted SARS-CoV-2 sequencing allowed for detection of important genetic mutations, such as spike: K417N, D614G, P681H, T716I, S982A, and D1118H, commonly associated with increased cell entry and reinfection. Microbiome analysis (DNA and RNA) provided important insight with respect to human health-related factors, including detection of pathogens and their virulence/antibiotic resistance genes. Specific microbial species comprising the wastewater microbiome correlated with incidence of SARS-CoV-2 RNA, suggesting potential association with SARS-CoV-2 infection. Climatic conditions, namely, temperature, were related to incidence of COVID-19 and detection of SARS-CoV-2 in wastewater, having been monitored as part of an environmental risk score assessment carried out in this study. In summary, the wastewater microbiome provides useful public health information, and hence, a valuable tool to proactively detect and characterize pathogenic agents circulating in a community. In effect, metagenomics of wastewater can serve as an early warning system for communicable diseases, by providing a larger source of information for health departments and public officials. IMPORTANCE Traditionally, testing for COVID-19 is done by detecting SARS-CoV-2 in samples collected from nasal swabs and/or saliva. However, SARS-CoV-2 can also be detected in feces of infected individuals. Therefore, wastewater samples can be used to test all individuals of a community contributing to the sewage collection system, i.e., the infrastructure, such as gravity pipes, manholes, tanks, lift stations, control structures, and force mains, that collects used water from residential and commercial sources and conveys the flow to a wastewater treatment plant. Here, we profile community wastewater collected from a manhole, detect presence of SARS-CoV-2, identify genetic mutations of SARS-CoV-2, and perform COVID-19 risk score assessment of the study area. Using metagenomics analysis, we also detect other microorganisms (bacteria, fungi, protists, and viruses) present in the samples. Results show that by analyzing all microorganisms present in wastewater, pathogens circulating in a community can provide an early warning for contagious diseases.
Glioblastoma multiforme (GBM) modulates the immune system to engance its malignant potential. Signal transducer and activator of transcription 3 (STAT3) activation is a regulatory node in modulating ...the immune microenvironment in several human tumors, including GBM. To investigate whether STAT3 inhibition might enhance anti-tumor responses, we inhibited STAT3 signaling using small interfering RNA against STAT3. We tested the human GBM cell lines U87, U251, and HS683, which are known to constitutively express high levels of phospho-STAT3. STAT3 inhibition resulted in enhanced expression of several pro-inflammatory cytokines and chemokines and supernatants from STAT3-silenced human GBM cell lines increased lipopolysaccharide-induced dendritic cell activation in vitro. We obtained comparable results when STAT3 activity was suppressed with specific small molecule inhibitors. Our results support the hypothesis that activated STAT3 contributes to the immunosuppressive microenvironment in GBM and support previous studies implicating STAT3 as a potential target for immunotherapy.
Regulatory T (Treg) cells suppress inflammatory immune responses and autoimmunity caused by self-reactive T cells. The key Treg cell transcription factor Foxp3 is downregulated during inflammation to ...allow for the acquisition of effector T cell-like functions. Here, we demonstrate that stress signals elicited by proinflammatory cytokines and lipopolysaccharides lead to the degradation of Foxp3 through the action of the E3 ubiquitin ligase Stub1. Stub1 interacted with Foxp3 to promote its K48-linked polyubiquitination in an Hsp70-dependent manner. Knockdown of endogenous Stub1 or Hsp70 prevented Foxp3 degradation. Furthermore, the overexpression of Stub1 in Treg cells abrogated their ability to suppress inflammatory immune responses in vitro and in vivo and conferred a T-helper-1-cell-like phenotype. Our results demonstrate the critical role of the stress-activated Stub1-Hsp70 complex in promoting Treg cell inactivation, thus providing a potential therapeutic target for the intervention against autoimmune disease, infection, and cancer.
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•Stress signals elicited by proinflammatory cytokines and LPS lead to loss of Foxp3•E3 ubiquitin ligase Stub1 interacts with and ubiquitinates Foxp3 for its degradation•Interaction between Stub1 and Foxp3 depends on the biochemical stress indicator Hsp70•Stub1 inhibits Treg function in vitro and in vivo
Abstract
CD4+ regulatory T cells (Tregs) maintain immunological self-tolerance and immune homeostasis by suppressing aberrant or excessive immune responses. The core genetic program of Tregs and ...their ability to suppress pathologic immune responses depends on the transcription factor Foxp3. Despite progress in understanding mechanisms of Foxp3-dependent gene activation, the molecular mechanism of Foxp3-dependent gene repression remains largely unknown. Here we identify Eos, a zinc-finger transcription factor of the Ikaros family, as a critical mediator of Foxp3-dependent gene silencing in Tregs. By qRT-PCR analysis we found that Eos is highly expressed in the CD4+CD25+ and CD4+Foxp3+ population of T cells. Additionally, Eos interacts directly with Foxp3 and induces chromatin modifications that result in gene silencing in Tregs. Silencing of Eos in Tregs abrogates their ability to suppress naïve T cell proliferation in vitro. Furthermore, Tregs with Eos knockdown fail to protect recipient Rag2-/- mice from severe colitis when co-transferred with naïve (CD4+CD25- CD62Lhigh ) T cells. Eos silencing in Tregs also endows these cells with partial effector function. These results demonstrate the critical role that Eos plays in Treg programming. They also raise the possibility that the interaction between Foxp3 and Eos may be a therapeutic target for controlling pathological immune responses in which excessive Treg activity plays an important role such as cancer and chronic infections.
T cell differentiation into distinct functional effector and inhibitory subsets is regulated, in part, by the cytokine environment present at the time of antigen recognition. Here, we show that ...hypoxia-inducible factor 1 (HIF-1), a key metabolic sensor, regulates the balance between regulatory T cell (T
reg) and T
H17 differentiation. HIF-1 enhances T
H17 development through direct transcriptional activation of RORγt and via tertiary complex formation with RORγt and p300 recruitment to the IL-17 promoter, thereby regulating T
H17 signature genes. Concurrently, HIF-1 attenuates T
reg development by binding Foxp3 and targeting it for proteasomal degradation. Importantly, this regulation occurs under both normoxic and hypoxic conditions. Mice with HIF-1α-deficient T cells are resistant to induction of T
H17-dependent experimental autoimmune encephalitis associated with diminished T
H17 and increased T
reg cells. These findings highlight the importance of metabolic cues in T cell fate determination and suggest that metabolic modulation could ameliorate certain T cell-based immune pathologies.
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► HIF-1 is induced by T
H17 promoting signals in a Stat3-dependent manner ► HIF-1 cooperates with RORγt, Stat3, and p300 to drive transcription of T
H17 genes ► HIF-1 negatively regulates Treg development by mediating Foxp3 protein degradation ► In vitro and in vivo T
H17 differentiation is deficient in T cells lacking HIF-1
The metabolic sensor HIF-1 enhances Th17 development while inhibiting Treg development, pointing to the importance of metabolic cues in determining T cell fate and setting the threshold for an inflammatory response.
T cell differentiation into distinct functional effector and inhibitory subsets is regulated, in part, by the cytokine environment present at the time of antigen recognition. Here, we show that ...hypoxia-inducible factor 1 (HIF-1), a key metabolic sensor, regulates the balance between regulatory T cell (T(reg)) and T(H)17 differentiation. HIF-1 enhances T(H)17 development through direct transcriptional activation of RORγt and via tertiary complex formation with RORγt and p300 recruitment to the IL-17 promoter, thereby regulating T(H)17 signature genes. Concurrently, HIF-1 attenuates T(reg) development by binding Foxp3 and targeting it for proteasomal degradation. Importantly, this regulation occurs under both normoxic and hypoxic conditions. Mice with HIF-1α-deficient T cells are resistant to induction of T(H)17-dependent experimental autoimmune encephalitis associated with diminished T(H)17 and increased T(reg) cells. These findings highlight the importance of metabolic cues in T cell fate determination and suggest that metabolic modulation could ameliorate certain T cell-based immune pathologies.
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