Deletion of adipocyte Sine Oculis Homeobox Homolog 1 prevents lipolysis and attenuates skin fibrosis.

Background: The cardinal feature of systemic sclerosis (SSc) is skin thickening and tightening. Targetable mechanisms for skin features remain elusive. Drugs successful in treating internal organ manifestations have failed efficacy in skin. Dermal white adipose tissue (DWAT) is amongst the understudied contributors to skin manifestations. This study proposes the role of sine oculis homeobox homolog 1 (SIX1), a gene previously unrecognized as a contributor to dermal lipoatrophy characteristic of early skin fibrosis in SSc. Methods: Skin gene expression of SIX1 was analyzed in the GENISOS and PRESS SSc cohorts. Correlation analysis was performed with Spearman rank analysis. Novel mouse models were developed using the Cre-loxp system to knock out Six1 in all cells and mature adipocytes. Subcutaneous bleomycin was used to model early DWAT atrophy and dermal fibrosis characteristic of SSc. Findings: SIX1 was upregulated in SSc skin, the expression of which correlates with adipose-associated genes and molecular pathways. Genetic deletion of Six1 in all cells in mice challenged with bleomycin abrogated end-stage fibrotic gene expression and dermal adipocyte shrinkage. Adipocyte specific Six1 deletion was able to attenuate the early increase in skin thickness, a hallmark of experimental skin fibrosis. Further studies revealed a link between elevated SIX1 and increased expression of SERPINE1 and its protein PAI-1 which are known pro-fibrotic mediators. Interpretation: This work identifies SIX1 as an early marker of skin fibrosis in SSc. We also demonstrate a causative role of Six1 in skin fibrosis by promoting adipocyte loss and show that deletion of Six1 in adipocytes has the potential of impacting early disease progression.

Hypoxia-adenosine axis as therapeutic targets for acute respiratory distress syndrome.

The human respiratory and circulatory systems collaborate intricately to ensure oxygen delivery to all cells, which is vital for ATP production and maintaining physiological functions and structures. During limited oxygen availability, hypoxia-inducible factors (HIFs) are stabilized and play a fundamental role in maintaining cellular processes for hypoxia adaptation. First discovered during investigations of erythropoietin production regulation, HIFs influence physiological and pathological processes, including development, inflammation, wound healing, and cancer. HIFs promote extracellular adenosine signaling by enhancing adenosine generation and receptor signaling, representing an endogenous feedback mechanism that curbs excessive inflammation, supports injury resolution, and enhances hypoxia tolerance. This is especially important for conditions that involve tissue hypoxia, such as acute respiratory distress syndrome (ARDS), which globally poses significant health challenges without specific treatment options. Consequently, pharmacological strategies to amplify HIF-mediated adenosine production and receptor signaling are of great importance.

Animal models in systemic sclerosis: an update.

PURPOSE OF REVIEW: Systemic sclerosis (SSc) is a multisystem autoimmune connective tissue disease characterized by early inflammation followed by excessive fibrosis in the skin and internal organs. Enhancing our comprehension of SSc pathogenesis is essential to develop effective therapeutic strategies. Animal models that mimic one or more aspects of SSc have been proven to be a valuable resource for investigating disease mechanisms. This review aims to provide an updated overview of the existing SSc animal models and the potentially relevant pathways to SSc pathogenesis. RECENT FINDINGS: This review focuses on the most recently generated and investigated animal models, which delve into novel pathways beyond existing models or employ genetic technologies to gain a deeper understanding of SSc pathogenesis including activation of early type I interferon (IFN) signaling pathway, immune cell function and pulmonary artery hypertension (PAH). SUMMARY: While no single animal model can fully replicate SSc, a combination of different models can offer valuable insights into the pathways involved in the onset and advancement of the SSc. These insights can prove animal models as a crutial preclinical tool for developing effective treatments for SSc.

Targeting myocardial equilibrative nucleoside transporter ENT1 provides cardioprotection by enhancing myeloid Adora2b signaling.

Previous studies implicate extracellular adenosine signaling in attenuating myocardial ischemia and reperfusion injury (IRI). This extracellular adenosine signaling is terminated by its uptake into cells by equilibrative nucleoside transporters (ENTs). Thus, we hypothesized that targeting ENTs would function to increase cardiac adenosine signaling and concomitant cardioprotection against IRI. Mice were exposed to myocardial ischemia and reperfusion injury. Myocardial injury was attenuated in mice treated with the nonspecific ENT inhibitor dipyridamole. A comparison of mice with global Ent1 or Ent2 deletion showed cardioprotection only in Ent1-/- mice. Moreover, studies with tissue-specific Ent deletion revealed that mice with myocyte-specific Ent1 deletion (Ent1loxP/loxP Myosin Cre+ mice) experienced smaller infarct sizes. Measurements of cardiac adenosine levels demonstrated that postischemic elevations of adenosine persisted during reperfusion after targeting ENTs. Finally, studies in mice with global or myeloid-specific deletion of the Adora2b adenosine receptor (Adora2bloxP/loxP LysM Cre+ mice) implied that Adora2b signaling on myeloid-inflammatory cells in cardioprotection provided by ENT inhibition. These studies reveal a previously unrecognized role for myocyte-specific ENT1 in cardioprotection by enhancing myeloid-dependent Adora2b signaling during reperfusion. Extension of these findings implicates adenosine transporter inhibitors in cardioprotection against ischemia and reperfusion injury.

Probiotic-Derived Ecto-5'-Nucleotidase Produces Anti-Inflammatory Adenosine Metabolites in Treg-Deficient Scurfy Mice.

Probiotic Limosilactobacillus reuteri DSM 17938 (DSM 17938) prolongs the survival of Treg-deficient scurfy (SF) mice and reduces multiorgan inflammation by a process requiring adenosine receptor 2A (A2A) on T cells. We hypothesized that L. reuteri-derived ecto-5'-nucleotidase (ecto-5'NT) activity acts to generate adenosine, which may be a central mediator for L. reuteri protection in SF mice. We evaluated DSM 17938-5'NT activity and the associated adenosine and inosine levels in plasma, gut, and liver of SF mice. We examined orally fed DSM 17938, DSM 17938Δ5NT (with a deleted 5'NT gene), and DSM 32846 (BG-R46) (a naturally selected strain derived from DSM 17938). Results showed that DSM 17938 and BG-R46 produced adenosine while "exhausting" AMP, whereas DSM 17938∆5NT did not generate adenosine in culture. Plasma 5'NT activity was increased by DSM 17938 or BG-R46, but not by DSM 17938Δ5NT in SF mice. BG-R46 increased both adenosine and inosine levels in the cecum of SF mice. DSM 17938 increased adenosine levels, whereas BG-R46 increased inosine levels in the liver. DSM 17938Δ5NT did not significantly change the levels of adenosine or inosine in the GI tract or the liver of SF mice. Although regulatory CD73+CD8+ T cells were decreased in spleen and blood of SF mice, these regulatory T cells could be increased by orally feeding DSM 17938 or BG-R46, but not DSM 17938Δ5NT. In conclusion, probiotic-5'NT may be a central mediator of DSM 17938 protection against autoimmunity. Optimal 5'NT activity from various probiotic strains could be beneficial in treating Treg-associated immune disorders in humans.

Probiotic-derived ecto-5'-nucleotidase produces anti-inflammatory adenosine metabolites in Treg-deficient scurfy mice.

Probiotic Limosilactobacillus reuteri DSM 17938 (DSM 17938) prolonges the survival of Treg-deficient scurfy (SF) mice and reduces multiorgan inflammation by a process requiring adenosine receptor 2A (A 2A ) on T cells. We hypothesized that L. reuteri -derived ecto-5'-nucleotidase (ecto-5'NT) activity acts to generate adenosine, which may be a central mediator for L. reuteri protection in SF mice. We evaluated DSM 17938-5'NT activity and the associated adenosine and inosine levels in plasma, gut and liver of SF mice. We examined orally fed DSM 17938, DSM 17938Δ5NT (with a deleted 5'NT gene), and DSM 32846 (BG-R46) (a naturally selected strain derived from DSM 17938). Results showed that DSM 17938 and BG-R46 produced adenosine while "exhausting" AMP, whereas DSM 17938∆5NT did not generate adenosine in culture. Plasma 5'NT activity was increased by DSM 17938 or BG-R46, but not by DSM 17938Δ5NT in SF mice. BG-R46 increased both adenosine and inosine levels in the cecum of SF mice. DSM 17938 increased adenosine levels, whereas BG-R46 increased inosine levels in the liver. DSM 17938Δ5NT did not significantly change the levels of adenosine or inosine in the GI tract or the liver of SF mice. Although regulatory CD73 + CD8 + T cells were decreased in spleen and blood of SF mice, these regulatory T cells could be increased by orally feeding DSM 17938 or BG-R46, but not DSM 17938Δ5NT. In conclusion, probiotic-5'NT may be a central mediator of DSM 17938 protection against autoimmunity. Optimal 5'NT activity from various probiotic strains could be beneficial in treating Treg-associated immune disorders in humans.

STAT6 suppression prevents bleomycin-induced dermal fibrosis.

Fibrosis of the skin and internal organs is a hallmark of systemic sclerosis (SSc). Although the pathogenesis of SSc is poorly understood, increasing evidence suggests that interleukins (IL)-4 and - 13 contribute to the pathogenesis of skin fibrosis by promoting collagen production and myofibroblast differentiation. Signal transducers and activators of transcription 6 (STAT6) is one of the most important downstream transcription factors activated by both IL-4 and IL-13. However, it is not completely understood whether STAT6 plays a role during the pathogenesis of skin fibrosis in SSc. In this study, we observed increased STAT6 phosphorylation in fibrotic skin samples collected from SSc patients as well as bleomycin-injected murine mice. Knockout of Stat6 in mice significantly (1) suppressed the expression of fibrotic cytokines including Il13, Il17, Il22, Ccl2, and the alternatively activated macrophage marker Cd206; (2) reduced the production of collagen and fibronectin, and (3) attenuated late-stage skin fibrosis and inflammation induced by bleomycin. Consistently, mice treated with STAT6 inhibitor AS1517499 also attenuated skin fibrosis on day 28. In addition, a co-culture experiment demonstrated that skin epithelial cells with STAT6 knockdown had reduced cytokine expression in response to IL-4/IL-13, and subsequently attenuated fibrotic protein expression in skin fibroblasts. On the other side, STAT6 depletion in skin fibroblasts attenuated IL-4/IL-13-induced cytokine and fibrotic marker expression, and reduced CXCL2 expression in co-cultured keratinocytes. In summary, our study highlighted an important yet not fully understood role of STAT6 in skin fibrosis by driving innate inflammation and differentiation of alternatively activated macrophages in response to injury.

CD73-Dependent Adenosine Signaling through Adora2b Drives Immunosuppression in Ductal Pancreatic Cancer.

The microenvironment that surrounds pancreatic ductal adenocarcinoma (PDAC) is profoundly desmoplastic and immunosuppressive. Understanding triggers of immunosuppression during the process of pancreatic tumorigenesis would aid in establishing targets for effective prevention and therapy. Here, we interrogated differential molecular mechanisms dependent on cell of origin and subtype that promote immunosuppression during PDAC initiation and in established tumors. Transcriptomic analysis of cell-of-origin-dependent epithelial gene signatures revealed that Nt5e/CD73, a cell-surface enzyme required for extracellular adenosine generation, is one of the top 10% of genes overexpressed in murine tumors arising from the ductal pancreatic epithelium as opposed to those rising from acinar cells. These findings were confirmed by IHC and high-performance liquid chromatography. Analysis in human PDAC subtypes indicated that high Nt5e in murine ductal PDAC models overlaps with high NT5E in human PDAC squamous and basal subtypes, considered to have the highest immunosuppression and worst prognosis. Multiplex immunofluorescent analysis showed that activated CD8+ T cells in the PDAC tumor microenvironment express high levels of CD73, indicating an opportunity for immunotherapeutic targeting. Delivery of CD73 small-molecule inhibitors through various delivery routes reduced tumor development and growth in genetically engineered and syngeneic mouse models. In addition, the adenosine receptor Adora2b was a determinant of adenosine-mediated immunosuppression in PDAC. These findings highlight a molecular trigger of the immunosuppressive PDAC microenvironment elevated in the ductal cell of origin, linking biology with subtype classification, critical components for PDAC immunoprevention and personalized approaches for immunotherapeutic intervention. SIGNIFICANCE: Ductal-derived pancreatic tumors have elevated epithelial and CD8+GZM+ T-cell CD73 expression that confers sensitivity to small-molecule inhibition of CD73 or Adora2b to promote CD8+ T-cell-mediated tumor regression. See related commentary by DelGiorno, p. 977.

CD73-generated extracellular adenosine promotes resolution of neutrophil-mediated tissue injury and restrains metaplasia in pancreatitis.

Pancreatitis is currently the leading cause of gastrointestinal hospitalizations in the US. This condition occurs in response to abdominal injury, gallstones, chronic alcohol consumption or, less frequently, the cause remains idiopathic. CD73 is a cell surface ecto-5'-nucleotidase that generates extracellular adenosine, which can contribute to resolution of inflammation by binding adenosine receptors on infiltrating immune cells. We hypothesized genetic deletion of CD73 would result in more severe pancreatitis due to decreased generation of extracellular adenosine. CD73 knockout (CD73-/- ) and C57BL/6 (wild type, WT) mice were used to evaluate the progression and response of caerulein-induced acute and chronic pancreatitis. In response to caerulein-mediated chronic or acute pancreatitis, WT mice display resolution of pancreatitis at earlier timepoints than CD73-/- mice. Using immunohistochemistry and analysis of single-cell RNA-seq (scRNA-seq) data, we determined CD73 localization in chronic pancreatitis is primarily observed in mucin/ductal cell populations and immune cells. In murine pancreata challenged with caerulein to induce acute pancreatitis, we compared CD73-/- to WT mice and observed a significant infiltration of Ly6G+, MPO+, and Granzyme B+ cells in CD73-/- compared to WT pancreata and we quantified a significant increase in acinar-to-ductal metaplasia demonstrating sustained metaplasia and inflammation in CD73-/- mice. Using neutrophil depletion in CD73-/- mice, we show neutrophil depletion significantly reduces metaplasia defined by CK19+ cells per field and significantly reduces acute pancreatitis. These data identify CD73 enhancers as a potential therapeutic strategy for patients with acute and chronic pancreatitis as adenosine generation and activation of adenosine receptors is critical to resolve persistent inflammation in the pancreas.

Radiofrequency ablation in combination with CD73 inhibitor AB680 reduces tumor growth and enhances anti-tumor immunity in a syngeneic model of pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma presents a 5-year overall survival rate of 11%, placing an imperative need for the discovery and application of innovative treatments. Radiofrequency ablation represents a promising therapy for PDA, as studies show it induces coagulative necrosis and a host adaptive immune response. In this work we evaluated the effects of RFA treatment in vivo by establishing a syngeneic mouse model of PDA and performing tumor ablation in one flank. Our studies revealed RFA acutely impaired PDA tumor growth; however, such effects were not sustained one week after treatment. Adenosine (ADO) pathway represents a strong immunosuppressive mechanism that was shown to play a role in PDA progression and preliminary data from ongoing clinical studies suggest ADO pathway inhibition may improve therapeutic outcomes. Thus, to investigate whether ADO generation may be involved in tumor growth relapse after RFA, we evaluated adenosine-monophosphate (AMP), ADO and inosine (INO) levels by HPLC and found they were acutely increased after treatment. Thus, we evaluated an in vivo CD73 inhibition in combination with RFA to study ADO pathway implication in RFA response. Results showed combination therapy of RFA and a CD73 small molecule inhibitor (AB680) in vivo promoted sustained tumor growth impairment up to 10 days after treatment as evidenced by increased necrosis and anti-tumor immunity, suggesting RFA in combination with CD73 inhibitors may improve PDA patient response.

Alternative adenosine Receptor activation: The netrin-Adora2b link.

During hypoxia or inflammation, extracellular adenosine levels are elevated. Studies using pharmacologic approaches or genetic animal models pertinent to extracellular adenosine signaling implicate this pathway in attenuating hypoxia-associated inflammation. There are four distinct adenosine receptors. Of these, it is not surprising that the Adora2b adenosine receptor functions as an endogenous feedback loop to control hypoxia-associated inflammation. First, Adora2b activation requires higher adenosine concentrations compared to other adenosine receptors, similar to those achieved during hypoxic inflammation. Second, Adora2b is transcriptionally induced during hypoxia or inflammation by hypoxia-inducible transcription factor HIF1A. Studies seeking an alternative adenosine receptor activation mechanism have linked netrin-1 with Adora2b. Netrin-1 was originally discovered as a neuronal guidance molecule but also functions as an immune-modulatory signaling molecule. Similar to Adora2b, netrin-1 is induced by HIF1A, and has been shown to enhance Adora2b signaling. Studies of acute respiratory distress syndrome (ARDS), intestinal inflammation, myocardial or hepatic ischemia and reperfusion implicate the netrin-Adora2b link in tissue protection. In this review, we will discuss the potential molecular linkage between netrin-1 and Adora2b, and explore studies demonstrating interactions between netrin-1 and Adora2b in attenuating tissue inflammation.

The Hypoxia-Adenosine Link during Myocardial Ischemia-Reperfusion Injury.

Despite increasing availability and more successful interventional approaches to restore coronary reperfusion, myocardial ischemia-reperfusion injury is a substantial cause of morbidity and mortality worldwide. During myocardial ischemia, the myocardium becomes profoundly hypoxic, thus causing stabilization of hypoxia-inducible transcription factors (HIF). Stabilization of HIF leads to a transcriptional program that promotes adaptation to hypoxia and cellular survival. Transcriptional consequences of HIF stabilization include increases in extracellular production and signaling effects of adenosine. Extracellular adenosine functions as a signaling molecule via the activation of adenosine receptors. Several studies implicated adenosine signaling in cardioprotection, particularly through the activation of the Adora2a and Adora2b receptors. Adenosine receptor activation can lead to metabolic adaptation to enhance ischemia tolerance or dampen myocardial reperfusion injury via signaling events on immune cells. Many studies highlight that clinical strategies to target the hypoxia-adenosine link could be considered for clinical trials. This could be achieved by using pharmacologic HIF activators or by directly enhancing extracellular adenosine production or signaling as a therapy for patients with acute myocardial infarction, or undergoing cardiac surgery.

SARS-CoV-2 Infection: Host Response, Immunity, and Therapeutic Targets.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has resulted in a global pandemic with severe socioeconomic effects. Immunopathogenesis of COVID-19 leads to acute respiratory distress syndrome (ARDS) and organ failure. Binding of SARS-CoV-2 spike protein to human angiotensin-converting enzyme 2 (hACE2) on bronchiolar and alveolar epithelial cells triggers host inflammatory pathways that lead to pathophysiological changes. Proinflammatory cytokines and type I interferon (IFN) signaling in alveolar epithelial cells counter barrier disruption, modulate host innate immune response to induce chemotaxis, and initiate the resolution of inflammation. Here, we discuss experimental models to study SARS-CoV-2 infection, molecular pathways involved in SARS-CoV-2-induced inflammation, and viral hijacking of anti-inflammatory pathways, such as delayed type-I IFN response. Mechanisms of alveolar adaptation to hypoxia, adenosinergic signaling, and regulatory microRNAs are discussed as potential therapeutic targets for COVID-19.

Ca2+-dependent protein acyltransferase DHHC21 controls activation of CD4+ T cells.

Despite the recognized significance of reversible protein lipidation (S-acylation) for T cell receptor signal transduction, the enzymatic control of this post-translational modification in T cells remains poorly understood. Here, we demonstrate that DHHC21 (also known as ZDHHC21), a member of the DHHC family of mammalian protein acyltransferases, mediates T cell receptor-induced S-acylation of proximal T cell signaling proteins. Using Zdhhc21dep mice, which express a functionally deficient version of DHHC21, we show that DHHC21 is a Ca2+/calmodulin-dependent enzyme critical for activation of naïve CD4+ T cells in response to T cell receptor stimulation. We find that disruption of the Ca2+/calmodulin-binding domain of DHHC21 does not affect thymic T cell development but prevents differentiation of peripheral CD4+ T cells into Th1, Th2 and Th17 effector T helper lineages. Our findings identify DHHC21 as an essential component of the T cell receptor signaling machinery and define a new role for protein acyltransferases in regulation of T cell-mediated immunity.

PMN-derived netrin-1 attenuates cardiac ischemia-reperfusion injury via myeloid ADORA2B signaling.

Previous studies implicated the neuronal guidance molecule netrin-1 in attenuating myocardial ischemia-reperfusion injury. However, the tissue-specific sources and receptor signaling events remain elusive. Neutrophils are among the first cells responding to an ischemic insult and can be associated with tissue injury or rescue. We found netrin-1 levels were elevated in the blood of patients with myocardial infarction, as well as in mice exposed to myocardial ischemia-reperfusion. Selectively increased infarct sizes and troponin levels were found in Ntn1loxP/loxP Lyz2 Cre+ mice, but not in mice with conditional netrin-1 deletion in other tissue compartments. In vivo studies using neutrophil depletion identified neutrophils as the main source for elevated blood netrin-1 during myocardial injury. Finally, pharmacologic studies using treatment with recombinant netrin-1 revealed a functional role for purinergic signaling events through the myeloid adenosine A2b receptor in mediating netrin-1-elicited cardioprotection. These findings suggest an autocrine signaling loop with a functional role for neutrophil-derived netrin-1 in attenuating myocardial ischemia-reperfusion injury through myeloid adenosine A2b signaling.

Hypoxia-inducible factor-dependent induction of myeloid-derived netrin-1 attenuates natural killer cell infiltration during endotoxin-induced lung injury.

Sepsis and sepsis-associated lung inflammation significantly contribute to the morbidity and mortality of critical illness. Here, we examined the hypothesis that neuronal guidance proteins could orchestrate inflammatory events during endotoxin-induced lung injury. Through a targeted array, we identified netrin-1 as the top upregulated neuronal guidance protein in macrophages treated with lipopolysaccharide (LPS). Furthermore, we found that netrin-1 is highly enriched in infiltrating myeloid cells, particularly in macrophages during LPS-induced lung injury. Transcriptional studies implicate hypoxia-inducible factor HIF-1α in the transcriptional induction of netrin-1 during LPS treatment. Subsequently, the deletion of netrin-1 in the myeloid compartment (Ntn1loxp/loxp LysM Cre) resulted in exaggerated mortality and lung inflammation. Surprisingly, further studies revealed enhanced natural killer cells (NK cells) infiltration in Ntn1loxp/loxp LysM Cre mice, and neutralization of NK cell chemoattractant chemokine (C-C motif) ligand 2 (CCL2) reversed the exaggerated lung inflammation. Together, these studies provide functional insight into myeloid cell-derived netrin-1 in controlling lung inflammation through the modulation of CCL2-dependent infiltration of NK cells.

Adenosine at the Interphase of Hypoxia and Inflammation in Lung Injury.

Hypoxia and inflammation often coincide in pathogenic conditions such as acute respiratory distress syndrome (ARDS) and chronic lung diseases, which are significant contributors to morbidity and mortality for the general population. For example, the recent global outbreak of Coronavirus disease 2019 (COVID-19) has placed viral infection-induced ARDS under the spotlight. Moreover, chronic lung disease ranks the third leading cause of death in the United States. Hypoxia signaling plays a diverse role in both acute and chronic lung inflammation, which could partially be explained by the divergent function of downstream target pathways such as adenosine signaling. Particularly, hypoxia signaling activates adenosine signaling to inhibit the inflammatory response in ARDS, while in chronic lung diseases, it promotes inflammation and tissue injury. In this review, we discuss the role of adenosine at the interphase of hypoxia and inflammation in ARDS and chronic lung diseases, as well as the current strategy for therapeutic targeting of the adenosine signaling pathway.

Transforming growth factor ß1 alters the 3'-UTR of mRNA to promote lung fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic disease characterized by the pathological remodeling of air sacs as a result of excessive accumulation of extracellular matrix (ECM) proteins, but the mechanism governing the robust protein expression is poorly understood. Our recent findings demonstrate that alternative polyadenylation (APA) caused by NUDT21 reduction is important for the increased expression of fibrotic mediators and ECM proteins in lung fibroblasts by shortening the 3'-untranslated regions (3'-UTRs) of mRNAs and stabilizing their transcripts, therefore activating pathological signaling pathways. Despite the importance of NUDT21 reduction in the regulation of fibrosis, the underlying mechanisms for the depletion are unknown. We demonstrate here that NUDT21 is depleted by TGFß1. We found that miR203, which is increased in IPF, was induced by TGFß1 to target the NUDT21 3'-UTR, thus depleting NUDT21 in human and mouse lung fibroblasts. TGFß1-mediated NUDT21 reduction was attenuated by the miR203 inhibitor antagomiR203 in fibroblasts. TGFß1 transgenic mice revealed that TGFß1 down-regulates NUDT21 in fibroblasts in vivo Furthermore, TGFß1 promoted differential APA of fibrotic genes, including FGF14, RICTOR, TMOD2, and UCP5, in association with increased protein expression. This unique differential APA signature was also observed in IPF fibroblasts. Altogether, our results identified TGFß1 as an APA regulator through NUDT21 depletion amplifying pulmonary fibrosis.

Comprehensive characterization of circular RNAs in ~ 1000 human cancer cell lines.

BACKGROUND: Human cancer cell lines are fundamental models for cancer research and therapeutic strategy development. However, there is no characterization of circular RNAs (circRNAs) in a large number of cancer cell lines. METHODS: Here, we apply four circRNA identification algorithms to heuristically characterize the expression landscape of circRNAs across ~ 1000 human cancer cell lines from CCLE polyA-enriched RNA-seq data. By using integrative analysis and experimental approaches, we explore the expression landscape, biogenesis, functional consequences, and drug response of circRNAs across different cancer lineages. RESULTS: We revealed highly lineage-specific expression patterns of circRNAs, suggesting that circRNAs may be powerful diagnostic and/or prognostic markers in cancer treatment. We also identified key genes involved in circRNA biogenesis and confirmed that TGF-ß signaling may promote biogenesis of circRNAs. Strikingly, we showed that clinically actionable genes are more likely to generate circRNAs, potentially due to the enrichment of RNA-binding protein (RBP) binding sites. Among these, circMYC can promote cell proliferation. We observed strong association between the expression of circRNAs and the response to drugs, especially those targeting chromatin histone acetylation. Finally, we developed a user-friendly data portal, CircRNAs in cancer cell lines (CircRiC, https://hanlab.uth.edu/cRic ), to benefit the biomedical research community. CONCLUSIONS: Our study provides the characterization of circRNAs in cancer cell lines and explored the potential mechanism of circRNA biogenesis as well as its therapeutic implications. We also provide a data portal to facilitate the related biomedical researches.