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Cell Reports May 2023Cellular metabolism is tightly regulated by growth factor signaling, which promotes metabolic rewiring to support growth and proliferation. While growth factor-induced...
Cellular metabolism is tightly regulated by growth factor signaling, which promotes metabolic rewiring to support growth and proliferation. While growth factor-induced transcriptional and post-translational modes of metabolic regulation have been well defined, whether post-transcriptional mechanisms impacting mRNA stability regulate this process is less clear. Here, we present the ZFP36/L1/L2 family of RNA-binding proteins and mRNA decay factors as key drivers of metabolic regulation downstream of acute growth factor signaling. We quantitatively catalog metabolic enzyme and nutrient transporter mRNAs directly bound by ZFP36 following growth factor stimulation-many of which encode rate-limiting steps in metabolic pathways. Further, we show that ZFP36 directly promotes the mRNA decay of Enolase 2 (Eno2), altering Eno2 protein expression and enzymatic activity, and provide evidence of a ZFP36/Eno2 axis during VEGF-stimulated developmental retinal angiogenesis. Thus, ZFP36-mediated mRNA decay serves as an important mode of metabolic regulation downstream of growth factor signaling within dynamic cell and tissue states.
Topics: RNA-Binding Proteins; Signal Transduction; Intercellular Signaling Peptides and Proteins; RNA Stability; Tristetraprolin
PubMed: 37086408
DOI: 10.1016/j.celrep.2023.112411 -
Proceedings of the National Academy of... Oct 2023Externalized histones erupt from the nucleus as extracellular traps, are associated with several acute and chronic lung disorders, but their implications in the...
Externalized histones erupt from the nucleus as extracellular traps, are associated with several acute and chronic lung disorders, but their implications in the molecular pathogenesis of interstitial lung disease are incompletely defined. To investigate the role and molecular mechanisms of externalized histones within the immunologic networks of pulmonary fibrosis, we studied externalized histones in human and animal bronchoalveolar lavage (BAL) samples of lung fibrosis. Neutralizing anti-histone antibodies were administered in bleomycin-induced fibrosis of C57BL/6 J mice, and subsequent studies used conditional/constitutive knockout mouse strains for TGFβ and IL-27 signaling along with isolated platelets and cultured macrophages. We found that externalized histones (citH3) were significantly ( < 0.01) increased in cell-free BAL fluids of patients with idiopathic pulmonary fibrosis (IPF; = 29) as compared to healthy controls ( = 10). The pulmonary sources of externalized histones were Ly6GCD11b neutrophils and nonhematopoietic cells after bleomycin in mice. Neutralizing monoclonal anti-histone H2A/H4 antibodies reduced the pulmonary collagen accumulation and hydroxyproline concentration. Histones activated platelets to release TGFβ1, which signaled through the TGFbRI/TGFbRII receptor complex on LysM cells to antagonize macrophage-derived IL-27 production. TGFβ1 evoked multiple downstream mechanisms in macrophages, including p38 MAPK, tristetraprolin, IL-10, and binding of SMAD3 to the IL-27 promotor regions. IL-27RA-deficient mice displayed more severe collagen depositions suggesting that intact IL-27 signaling limits fibrosis. In conclusion, externalized histones inactivate a safety switch of antifibrotic, macrophage-derived IL-27 by boosting platelet-derived TGFβ1. Externalized histones are accessible to neutralizing antibodies for improving the severity of experimental pulmonary fibrosis.
Topics: Humans; Mice; Animals; Mice, Inbred C57BL; Histones; Interleukin-27; Blood Platelets; Idiopathic Pulmonary Fibrosis
PubMed: 37756334
DOI: 10.1073/pnas.2215421120 -
Inflammation and Regeneration Mar 2024Inflammation orchestrates a finely balanced process crucial for microorganism elimination and tissue injury protection. A multitude of immune and non-immune cells,... (Review)
Review
Inflammation orchestrates a finely balanced process crucial for microorganism elimination and tissue injury protection. A multitude of immune and non-immune cells, alongside various proinflammatory cytokines and chemokines, collectively regulate this response. Central to this regulation is post-transcriptional control, governing gene expression at the mRNA level. RNA-binding proteins such as tristetraprolin, Roquin, and the Regnase family, along with RNA modifications, intricately dictate the mRNA decay of pivotal mediators and regulators in the inflammatory response. Dysregulated activity of these factors has been implicated in numerous human inflammatory diseases, underscoring the significance of post-transcriptional regulation. The increasing focus on targeting these mechanisms presents a promising therapeutic strategy for inflammatory and autoimmune diseases. This review offers an extensive overview of post-transcriptional regulation mechanisms during inflammatory responses, delving into recent advancements, their implications in human diseases, and the strides made in therapeutic exploitation.
PubMed: 38491500
DOI: 10.1186/s41232-024-00326-5 -
Genes & Development Aug 2023Histidine (His) residues are methylated in various proteins, but their roles and regulation mechanisms remain unknown. Here, we show that carnosine N-methyltransferase 1...
Histidine (His) residues are methylated in various proteins, but their roles and regulation mechanisms remain unknown. Here, we show that carnosine N-methyltransferase 1 (CARNMT1), a known His methyltransferase of dipeptide carnosine (βAla-His), is a major His N1-position-specific methyltransferase. We found that 52 His sites in 20 proteins underwent CARNMT1-mediated methylation. The consensus methylation site for CARNMT1 was identified as Cx(F/Y)xH, a C3H zinc finger (C3H ZF) motif. CARNMT1-deficient and catalytically inactive mutant mice showed embryonic lethality. Among the CARNMT1 target C3H ZF proteins, RNA degradation mediated by Roquin and tristetraprolin (TTP) was affected by CARNMT1 and its enzymatic activity. Furthermore, the recognition of the 3' splice site of the CARNMT1 target C3H ZF protein U2AF1 was perturbed, and pre-mRNA alternative splicing (AS) was affected by CARNMT1 deficiency. These findings indicate that CARNMT1-mediated protein His methylation, which is essential for embryogenesis, plays roles in diverse aspects of RNA metabolism by targeting C3H ZF-type RNA-binding proteins and modulating their functions, including pre-mRNA AS and mRNA degradation regulation.
Topics: Animals; Mice; Mice, Inbred C3H; Carnosine; Histidine; RNA Precursors; Methyltransferases; RNA Splice Sites; Zinc Fingers
PubMed: 37612136
DOI: 10.1101/gad.350755.123 -
Pharmacological Research Aug 2023The p38-MK2 signaling axis functions as an initiator of inflammation. Targeting the p38-MK2 signaling axis represents a direct therapeutic intervention of inflammatory...
The p38-MK2 signaling axis functions as an initiator of inflammation. Targeting the p38-MK2 signaling axis represents a direct therapeutic intervention of inflammatory diseases. We described here a novel role of andrographolide (AG), a small-molecule ent-labdane natural compound, as an inhibitor of p38-MK2 axis via MK2 degradation. AG was found to bind to the activation loop of MK2, located at the interface of the p38-MK2 biomolecular complex. This interaction disrupted the complex formation and predisposed MK2 to proteasome-mediated degradation. We showed that AG induced MK2 degradation in a concentration- and time-dependent manner and exerted its anti-inflammatory effects by enhancing the mRNA-destabilizing activity of tristetraprolin, thereby inhibiting pro-inflammatory mediator production (e.g., TNF-α, MCP-1). Administration of AG via intratracheal (i.t.) route to mice induced MK2 downregulation in lung alveolar macrophages, but not lung tissues, and prevented macrophage activation. Our study also demonstrated that the anti-inflammatory effects achieved by AG via MK2 degradation were more durable and sustained than that achieved by the conventional MK2 kinase inhibitors (e.g., PF-3644022). Taken together, our findings illustrated a novel mode of action of AG by modulating the p38-MK2 signaling axis and would pave the way for the development of a novel class of anti-inflammatory agents targeting MK2 for degradation by harnessing the privileged scaffold of AG.
Topics: Mice; Animals; Protein Serine-Threonine Kinases; Intracellular Signaling Peptides and Proteins; Anti-Inflammatory Agents; Diterpenes; p38 Mitogen-Activated Protein Kinases
PubMed: 37480973
DOI: 10.1016/j.phrs.2023.106861 -
Cell Death & Disease Aug 2023Non-small cell lung cancer (NSCLC) is the most common type of lung cancer, with high morbidity and mortality worldwide. Although the dysregulation of BARX1 expression...
Non-small cell lung cancer (NSCLC) is the most common type of lung cancer, with high morbidity and mortality worldwide. Although the dysregulation of BARX1 expression has been shown to be associated with malignant cancers, including NSCLC, the underlying mechanism remains elusive. In this study, we identified BARX1 as a common differentially expressed gene in lung squamous cell carcinoma and adenocarcinoma. Importantly, we uncovered a novel mechanism behind the regulation of BARX1, in which ZFP36 interacted with 3'UTR of BARX1 mRNA to mediate its destabilization. Loss of ZFP36 led to the upregulation of BARX1, which further promoted the proliferation, migration and invasion of NSCLC cells. In addition, the knockdown of BARX1 inhibited tumorigenicity in mouse xenograft. We demonstrated that BARX1 promoted the malignant phenotypes by transactivating a set of master oncogenes involved in the cell cycle, DNA synthesis and metastasis. Overall, our study provides insights into the mechanism of BARX1 actions in NSCLC and aids a better understanding of NSCLC pathogenesis.
Topics: Animals; Humans; Mice; Adenocarcinoma; Carcinoma, Non-Small-Cell Lung; Homeodomain Proteins; Lung Neoplasms; Oncogenes; Phenotype; Transcription Factors; Tristetraprolin
PubMed: 37587140
DOI: 10.1038/s41419-023-06044-z -
MBio Oct 2023Our findings define a novel role for ZIKV-induced TTP expression in regulating IFNβ/IFNλ production in primary hBMECs and Sertoli cells. These cells comprise key...
Our findings define a novel role for ZIKV-induced TTP expression in regulating IFNβ/IFNλ production in primary hBMECs and Sertoli cells. These cells comprise key physiological barriers subverted by ZIKV to access brain and testicular compartments and serve as reservoirs for persistent replication and dissemination. We demonstrate for the first time that the ARE-binding protein TTP is virally induced and post-transcriptionally regulates IFNβ/IFNλ secretion. In ZIKV-infected hBMEC and Sertoli cells, TTP knockout increased IFNβ/IFNλ secretion, while TTP expression blocked IFNβ/IFNλ secretion. The TTP-directed blockade of IFN secretion permits ZIKV spread and persistence in hBMECs and Sertoli cells and may similarly augment ZIKV spread across IFNλ-protected placental barriers. Our work highlights the importance of post-transcriptional ZIKV regulation of IFN expression and secretion in cells that regulate viral access to protected compartments and defines a novel mechanism of ZIKV-regulated IFN responses which may facilitate neurovirulence and sexual transmission.
Topics: Pregnancy; Male; Female; Humans; Sertoli Cells; Zika Virus; Zika Virus Infection; Tristetraprolin; Placenta; Virus Replication
PubMed: 37707056
DOI: 10.1128/mbio.01742-23 -
Frontiers in Cell and Developmental... 2023Asthma is the most common chronic inflammatory disease of the airways. The airway epithelium is a key driver of the disease, and numerous studies have established...
Asthma is the most common chronic inflammatory disease of the airways. The airway epithelium is a key driver of the disease, and numerous studies have established genome-wide differences in mRNA expression between health and asthma. However, the underlying molecular mechanisms for such differences remain poorly understood. The human TTP family is comprised of ZFP36, ZFP36L1 and ZFP36L2, and has essential roles in immune regulation by determining the stability and translation of myriad mRNAs encoding for inflammatory mediators. We investigated the expression and possible role of the tristetraprolin (TTP) family of RNA binding proteins (RBPs), poorly understood in asthma. We analysed the levels of , and mRNA in several publicly available asthma datasets, including single cell RNA-sequencing. We also interrogated the expression of known targets of these RBPs in asthma. We assessed the lung mRNA expression and cellular localization of Zfp36l1 and Zfp36l2 in precision cut lung slices in murine asthma models. Finally, we determined the expression in airway epithelium of ZFP36L1 and ZFP36L2 in human bronchial biopsies and performed rescue experiments in primary bronchial epithelium from patients with severe asthma. We found and mRNA levels significantly downregulated in the airway epithelium of patients with very severe asthma in different cohorts (5 healthy vs. 8 severe asthma; 36 moderate asthma vs. 37 severe asthma on inhaled steroids vs. 26 severe asthma on oral corticoids). Integrating several datasets allowed us to infer that mRNAs potentially targeted by these RBPs are increased in severe asthma. Zfp36l1 was downregulated in the lung of a mouse model of asthma, and immunostaining of lung slices with a dual antibody demonstrated that Zfp36l1/l2 nuclear localization was increased in the airway epithelium of an acute asthma mouse model, which was further enhanced in a chronic model. Immunostaining of human bronchial biopsies showed that airway epithelial cell staining of ZFP36L1 was decreased in severe asthma as compared with mild, while ZFP36L2 was upregulated. Restoring the levels of ZFP36L1 and ZFP36L2 in primary bronchial epithelial cells from patients with severe asthma decreased the mRNA expression of , and . We propose that the dysregulation of ZFP36L1/L2 levels as well as their subcellular mislocalization contributes to changes in mRNA expression and cytoplasmic fate in asthma.
PubMed: 37928904
DOI: 10.3389/fcell.2023.1241008 -
Cell Reports Jan 2024Vascular morphogenesis requires a delicate gradient of Notch signaling controlled, in part, by the distribution of ligands (Dll4 and Jagged1). How Jagged1 (JAG1)...
Vascular morphogenesis requires a delicate gradient of Notch signaling controlled, in part, by the distribution of ligands (Dll4 and Jagged1). How Jagged1 (JAG1) expression is compartmentalized in the vascular plexus remains unclear. Here, we show that Jag1 mRNA is a direct target of zinc-finger protein 36 (ZFP36), an RNA-binding protein involved in mRNA decay that we find robustly induced by vascular endothelial growth factor (VEGF). Endothelial cells lacking ZFP36 display high levels of JAG1 and increase angiogenic sprouting in vitro. Furthermore, mice lacking Zfp36 in endothelial cells display mispatterned and increased levels of JAG1 in the developing retinal vascular plexus. Abnormal levels of JAG1 at the sprouting front alters NOTCH1 signaling, increasing the number of tip cells, a phenotype that is rescued by imposing haploinsufficiency of Jag1. Our findings reveal an important feedforward loop whereby VEGF stimulates ZFP36, consequently suppressing Jag1 to enable adequate levels of Notch signaling during sprouting angiogenesis.
Topics: Animals; Mice; Calcium-Binding Proteins; Cell Proliferation; Endothelial Cells; Jagged-1 Protein; Membrane Proteins; Neovascularization, Physiologic; Receptors, Notch; Signal Transduction; Vascular Endothelial Growth Factor A
PubMed: 38157296
DOI: 10.1016/j.celrep.2023.113627 -
Scientific Reports Feb 2024Loss of inflammatory effector function, such as cytokine production and proliferation, is a fundamental driver of failure in T cell therapies against solid tumors. Here,...
Loss of inflammatory effector function, such as cytokine production and proliferation, is a fundamental driver of failure in T cell therapies against solid tumors. Here, we used CRISPR/Cas9 to genetically disrupt ZFP36, an RNA binding protein that regulates the stability of mRNAs involved in T cell inflammatory function, such as the cytokines IL2 and IFNγ, in human T cells engineered with a clinical-stage mesothelin-targeting CAR to determine whether its disruption could enhance antitumor responses. ZFP36 disruption slightly increased antigen-independent activation and cytokine responses but did not enhance overall performance in vitro or in vivo in a xenograft tumor model with NSG mice. While ZFP36 disruption does not reduce the function of CAR-T cells, these results suggest that singular disruption of ZFP36 is not sufficient to improve their function and may benefit from a multiplexed approach.
Topics: Humans; Animals; Mice; Mesothelin; Immunotherapy, Adoptive; T-Lymphocytes; Immunity; Cytokines; Disease Models, Animal; Xenograft Model Antitumor Assays; Cell Line, Tumor; Tristetraprolin
PubMed: 38326511
DOI: 10.1038/s41598-024-53769-8