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1.
Nat Commun ; 11(1): 2286, 2020 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385332

RESUMO

Studies on macrophage gene expression have historically focused on events leading to RNA polymerase II recruitment and transcription initiation, whereas the contribution of post-initiation steps to macrophage activation remains poorly understood. Here, we report that widespread promoter-proximal RNA polymerase II pausing in resting macrophages is marked by co-localization of the negative elongation factor (NELF) complex and facilitated by PU.1. Upon inflammatory stimulation, over 60% of activated transcriptome is regulated by polymerase pause-release and a transient genome-wide NELF dissociation from chromatin, unexpectedly, independent of CDK9, a presumed NELF kinase. Genetic disruption of NELF in macrophages enhanced transcription of AP-1-encoding Fos and Jun and, consequently, AP-1 targets including Il10. Augmented expression of IL-10, a critical anti-inflammatory cytokine, in turn, attenuated production of pro-inflammatory mediators and, ultimately, macrophage-mediated inflammation in vivo. Together, these findings establish a previously unappreciated role of NELF in constraining transcription of inflammation inhibitors thereby enabling inflammatory macrophage activation.

2.
Artigo em Inglês | MEDLINE | ID: mdl-32409604

RESUMO

Apicomplexan parasites use a specialized cilium structure called the apical complex to organize their secretory organelles and invasion machinery. The apical complex is integrally associated with both the parasite plasma membrane and an intermediate filament cytoskeleton called the inner-membrane complex (IMC). While the apical complex is essential to the parasitic lifestyle, little is known about the regulation of apical complex biogenesis. Here, we identify AC9 (apical cap protein 9), a largely intrinsically disordered component of the Toxoplasma gondii IMC, as essential for apical complex development, and therefore for host cell invasion and egress. Parasites lacking AC9 fail to successfully assemble the tubulin-rich core of their apical complex, called the conoid. We use proximity biotinylation to identify the AC9 interaction network, which includes the kinase extracellular signal-regulated kinase 7 (ERK7). Like AC9, ERK7 is required for apical complex biogenesis. We demonstrate that AC9 directly binds ERK7 through a conserved C-terminal motif and that this interaction is essential for ERK7 localization and function at the apical cap. The crystal structure of the ERK7-AC9 complex reveals that AC9 is not only a scaffold but also inhibits ERK7 through an unusual set of contacts that displaces nucleotide from the kinase active site. ERK7 is an ancient and autoactivating member of the mitogen-activated kinase (MAPK) family and its regulation is poorly understood in all organisms. We propose that AC9 dually regulates ERK7 by scaffolding and concentrating it at its site of action while maintaining it in an "off" state until the specific binding of a true substrate.

3.
Biopreserv Biobank ; 2020 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-32315536

RESUMO

Cryopreservation of red blood cells (RBCs) has been studied as a typical example of cryobiology methodology. To date, a mature and long-term cryopreservation process for RBCs has been developed, which has the weakness of complicated procedures due to high concentrations of glycerol (Gly). Therefore, it is still a research focus to find a new method for cryopreservation of RBCs to reduce the concentrations of cryoprotectants (CPAs). In this study, alginate hydrogels, which have been widely used in preservation research, were selected because of their advantages such as lower cost, cytocompatibility, and crosslinking occurring under mild conditions. With a variety of CPA solutions, the RBC recovery with the cryopreservation of RBCs and RBC-laden hydrogel microfibers was compared in different situations. It was found that hydrogel microfiber encapsulation enhances cryopreservation of human RBCs with low concentrations of CPAs. And Gly can be removed by washing directly with a 0.9% sodium chloride solution. This study validated the feasibility of cryopreservation of RBC-laden hydrogel microfibers. It may provide a new and evolving direction for reducing the concentrations of CPAs used in the preservation of RBCs.

4.
Phys Chem Chem Phys ; 2020 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-32301460

RESUMO

The traditional methods to predict electrokinetic energy conversion (EKEC) in nanochannels are mostly based on the Navier-Stokes (NS) equation for ionic flow and the Poisson-Boltzmann (PB) equation for charge distributions, which is questionable for ion transport through highly charged nanochannels. In this work, the classical density functional theory (cDFT) is used together with molecular dynamics (MD) simulation and the Navier-Stokes (NS) equation to predict the electrical current and the thermodynamic efficiency of electrokinetic energy conversion in nanochannels. By introducing numerical results for the slip length calculated from MD simulation, a significant increase of the electrokinetic current is predicted in comparison to that obtained from the traditional electrokinetic equations with the non-slip boundary condition, leading to the theoretical predictions of the thermodynamic efficiency for electrokinetic energy conversion in nanochannels in good agreement with recent experiments. The hybrid method predicts that maximum electrokinetic efficiency can be achieved by tuning the channel height and solution conditions including electrolyte concentrations, ion valences, and surface energies. The theoretical results provide new insights into pressure-driven electrical energy generation processes and helpful guidelines for engineering design and optimization of electrokinetic energy conversion.

5.
Food Funct ; 11(4): 3695-3705, 2020 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-32307472

RESUMO

Endometritis, the inflammation of the endometrial lining caused by bacterial pathogens, is associated with reproductive failure. Recent studies have shown that gut microbiota play an important role in infectious diseases. However, the roles of the gut microbiota in endometritis remain unclear. Here, we assessed the effects and mechanisms of the gut microbiota during endometritis induced by Staphylococcus aureus (S. aureus). A mouse gut microbiota-dysbiosis model was established by a mixture of antibiotics (Abx) and subsequently, a model of endometritis was established by the uterine perfusion of S. aureus. Fecal microbiota transplantation (FMT) was performed to evaluate the relationship between gut microbiota and endometritis. The results showed that the mice with gut microbiota-dysbiosis developed uterine inflammation, while this inflammatory response of the uterus was alleviated in mice with FMT to gut microbiota-dysbiosis. In addition, S. aureus-induced endometritis was greater in severity in the mice with gut dysbiosis as compared to the untreated mice. Moreover, these effects were reversed in mice with FMT to the gut microbiota-dysbiosis. GC-MS analysis demonstrated that the levels of short-chain fatty acids (SCFAs) in the feces of mice with gut microbiota-dysbiosis significantly decreased and pretreatment with sodium butyrate or sodium propionate increased the concentrations of butyrate or propionate in both the circulation and uterine tissues, thereby reducing the severity of endometritis induced by S. aureus. In addition, the increased pathogen load in the uteri of the mice with gut microbiota-dysbiosis was associated with a reduction in the phagocytic ability and responsiveness of neutrophils. In conclusion, the gut microbiota offer a protective effect against S. aureus-induced endometritis by regulating the levels of SCFAs and maintaining the phagocytic ability and responsiveness of neutrophils.

6.
ISME J ; 2020 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-32341472

RESUMO

Mastitis is one of the most prevalent diseases in dairy farming worldwide. The gut microbiota plays an important role in the regulation of systemic and local inflammatory diseases, such as mastitis. However, the regulatory mechanism of the gut microbiota on mastitis is still unclear. Thus, the aim of this study was to investigate the function and regulatory mechanisms of the gut microbiota in host defense against mastitis caused by Staphylococcus aureus (S. aureus) infection. Increased blood-milk barrier permeability, and S. aureus-induced mastitis severity were observed gut microbiota-dysbiosis mice compared with those in control mice. Moreover, feces microbiota transplantation (FMT) to microbbiota-dysbiosis mice reversed these effects. Furthermore, established disruption of commensal homeostasis results in significantly increased abundance of pathogenic Enterobacter bacteria, while the relative abundance of short-chain fatty acid (SCFAs)-producing bacterial phyla (Firmicutes and Bacteroidetes) was significantly reduced. However, FMT to gut microbiota-dysbiosis mice reversed these changes. In addition, dysbiosis reduced the levels of SCFAs, and administration of sodium propionate, sodium butyrate, and probiotics (butyrate-producing bacteria) reversed the changes in the blood-milk barrier and reduced the severity of mastitis induced by S. aureus. In conclusion, this new finding demonstrated that the gut microbiota acts as a protective factor in host defense against mastitis and that targeting the gut-mammary gland axis represents a promising therapeutic approach for mastitis treatment.

8.
Artigo em Inglês | MEDLINE | ID: mdl-32133747

RESUMO

Nanospaces are ubiquitous in the realm of biological systems and are of significant interest among supramolecular chemists. Understanding chemical behavior within nanospaces offers new perspectives on biological phenomena in Nature and opens the way to highly unusual and selective forms of catalysis. Supramolecular chemistry exploits weak, yet effective, intermolecular interactions such as hydrogen bonding, metal-ligand coordination, and the hydrophobic effect to assemble nano-sized molecular architectures, providing reactions with remarkable rate acceleration, substrate specificity, and product selectivity. In this minireview, we will focus on the strategies that supramolecular chemists use to emulate the efficiency of biological processes, and elucidate how chemical reactivity is efficiently controlled within well-defined nanospaces. Approaches such as orientation and proximity of substrate, transition state stabilization, and active site incorporation will be discussed.

9.
Adv Immunol ; 145: 129-157, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32081196

RESUMO

Immune responses are often accompanied by radical changes of cellular metabolism of immune cells. On the other hand, an ever increasing number of metabolic pathways and products have been found to possess immune regulatory functions. The field of immunometabolism that investigates the interplay between metabolism and immunity has developed rapidly during the past decade. In this chapter, we attempt to summarize the recent progresses by scientists in China on metabolic regulation of innate immunity from the following three perspectives: metabolic regulation of myeloid cell functions, metabolic adaptations of tissue resident myeloid cells, and metabolism and immunity at the mucosal surfaces.

11.
Cell Mol Immunol ; 2020 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-32015501

RESUMO

MicroRNAs (miRNAs) have been widely implicated in immune regulation, but evidence for the coordinated function of paralogous miRNA clusters remains scarce. Here, by using genetically modified mice with individual or combined cluster deficiencies, we found that three paralogous clusters of the miR-17~92 family of miRNAs collectively suppressed IL-12 production in macrophages. Accordingly, miR-17~92 family miRNAs deficiencies resulted in heightened production of IL-12 and thus enhanced the host defense against intracellular pathogen Listeria monocytogenes in vivo. Mechanistically, different members of the miR-17~92 family of miRNAs acted on a common target, PTEN, to inhibit IL-12 expression by modulating the PI3K-Akt-GSK3 pathway. In addition, the expression of miR-17~92 family miRNAs was collectively inhibited by the transcription factor RBP-J, and RBP-J-associated macrophage functional defects were genetically rescued by deleting three clusters of miR-17~92 family miRNAs on a RBP-J null background. Thus, our results illustrated key roles of three clusters of miR-17~92 family miRNAs in cooperatively controlling IL-12-mediated immune responses and identified miR-17~92 family miRNAs as functional targets of RBP-J in macrophages.

12.
Mol Biol Cell ; 31(9): 881-888, 2020 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-32073987

RESUMO

Primary cilia are important organizing centers that control diverse cellular processes. Apicomplexan parasites like Toxoplasma gondii have a specialized cilium-like structure called the conoid that organizes the secretory and invasion machinery critical for the parasites' lifestyle. The proteins that initiate the biogenesis of this structure are largely unknown. We identified the Toxoplasma orthologue of the conserved kinase ERK7 as essential to conoid assembly. Parasites in which ERK7 has been depleted lose their conoids late during maturation and are immotile and thus unable to invade new host cells. This is the most severe phenotype to conoid biogenesis yet reported, and is made more striking by the fact that ERK7 is not a conoid protein, as it localizes just basal to the structure. ERK7 has been recently implicated in ciliogenesis in metazoan cells, and our data suggest that this kinase has an ancient and central role in regulating ciliogenesis throughout Eukaryota.

13.
FASEB J ; 34(2): 2392-2407, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31908034

RESUMO

Bone resorption is a severe consequence of inflammatory diseases associated with osteolysis, such as rheumatoid arthritis (RA), often leading to disability in patients. In physiological conditions, the differentiation of bone-resorbing osteoclasts is delicately regulated by the balance between osteoclastogenic and anti-osteoclastogenic mechanisms. Inflammation has complex impact on osteoclastogenesis and bone destruction, and the underlying mechanisms of which, especially feedback inhibition, are underexplored. Here, we identify a novel regulatory network mediated by RBP-J/NFATc1-miR182 in TNF-induced osteoclastogenesis and inflammatory bone resorption. This network includes negative regulator RBP-J and positive regulators, NFATc1 and miR182, of osteoclast differentiation. In this network, miR182 is a direct target of both RBP-J and NFATc1. RBP-J represses, while NFATc1 activates miR182 expression through binding to specific open chromatin regions in the miR182 promoter. Inhibition of miR182 by RBP-J servers as a critical mechanism that limits TNF-induced osteoclast differentiation and inflammatory bone resorption. Inflammation, such as that which occurs in RA, shifts the expression levels of the components in this network mediated by RBP-J/NFATc1-miR182-FoxO3/PKR (previously identified miR182 targets) towards more osteoclastogenic, rather than healthy conditions. Treatment with TNF inhibitors in RA patients reverses the expression changes of the network components and osteoclastogenic potential. Thus, this network controls the balance between activating and repressive signals that determine the extent of osteoclastogenesis. These findings collectively highlight the biological significance and translational implication of this newly identified intrinsic regulatory network in inflammatory osteoclastogenesis and osteolysis.

14.
Cell Signal ; 69: 109549, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31987780

RESUMO

BACKGROUND: Our previous study showed that human omental adipose-derived stem cells (ADSCs) promote ovarian cancer growth and metastasis. In this study, the role of autophagy in the ovarian cancer-promoting effects of omental ADSCs was further determined. METHODS: The growth and invasion of ovarian cancer cells were detected by CCK-8 and Transwell assays, respectively. The autophagy of ovarian cancer cells transfected with MRFP-GFP-LC3 adenoviral vectors was evaluated by confocal microscopy and western blot assay. Transfection of STAT3 siRNA was used to inhibit the expression of STAT3. RESULTS: Our results show that autophagy plays a vital role in ovarian cancer and is promoted by ADSCs. Specifically, we show that proliferation and invasion are correlated with autophagy induction by ADSCs in two ovarian cancer cell lines under hypoxic conditions. Mechanistically, ADSCs activate the STAT3 signalling pathway, thereby promoting autophagy. Knockdown of STAT3 expression using siRNA decreased hypoxia-induced autophagy and decreased the proliferation and metastasis of ovarian cancer cells. CONCLUSION: Taken together, our data indicate that STAT3-mediated autophagy induced by ADSCs promotes ovarian cancer growth and metastasis.

15.
J Exp Med ; 217(4)2020 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-31944217

RESUMO

Macrophages play pleiotropic roles in maintaining the balance between immune tolerance and inflammatory responses in the gut. Here, we identified transcription factor RBP-J as a crucial regulator of colonic macrophage-mediated immune responses against the enteric pathogen Citrobacter rodentium. In the immune response phase, RBP-J promoted pathogen clearance by enhancing intestinal macrophage-elicited Th17 cell immune responses, which was achieved by maintenance of C/EBPß-dependent IL-6 production by overcoming miRNA-17∼92-mediated suppressive effects. RBP-J deficiency-associated phenotypes could be genetically corrected by further deleting miRNA-17∼92 in macrophages. In the late phase, noneradicated pathogens in RBP-J KO mice recruited abundant IL-1ß-expressing CD64+Ly6C+ colonic macrophages and thereby promoted persistence of ILC3-derived IL-22 to compensate for the impaired innate and adaptive immune responses, leading to ultimate clearance of pathogens. These results demonstrated that colonic macrophage-intrinsic RBP-J dynamically orchestrates intestinal immunity against pathogen infections by interfacing with key immune cells of T and innate lymphoid cell lineages.

17.
Protein Cell ; 11(2): 108-123, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31691194

RESUMO

Common γ chain cytokines are important for immune memory formation. Among them, the role of IL-2 remains to be fully explored. It has been suggested that this cytokine is critically needed in the late phase of primary CD4 T cell activation. Lack of IL-2 at this stage sets for a diminished recall response in subsequent challenges. However, as IL-2 peak production is over at this point, the source and the exact mechanism that promotes its production remain elusive. We report here that resting, previously antigen-stimulated CD4 T cells maintain a minimalist response to dendritic cells after their peak activation in vitro. This subtle activation event may be induced by DCs without overt presence of antigen and appears to be stronger if IL-2 comes from the same dendritic cells. This encounter reactivates a miniature IL-2 production and leads a gene expression profile change in these previously activated CD4 T cells. The CD4 T cells so experienced show enhanced reactivation intensity upon secondary challenges later on. Although mostly relying on in vitro evidence, our work may implicate a subtle programing for CD4 T cell survival after primary activation in vivo.

18.
Proc Natl Acad Sci U S A ; 116(51): 26008-26019, 2019 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-31796582

RESUMO

The transient receptor potential ankyrin 1 (TRPA1) channel functions as an irritant sensor and is a therapeutic target for treating pain, itch, and respiratory diseases. As a ligand-gated channel, TRPA1 can be activated by electrophilic compounds such as allyl isothiocyanate (AITC) through covalent modification or activated by noncovalent agonists through ligand binding. However, how covalent modification leads to channel opening and, importantly, how noncovalent binding activates TRPA1 are not well-understood. Here we report a class of piperidine carboxamides (PIPCs) as potent, noncovalent agonists of human TRPA1. Based on their species-specific effects on human and rat channels, we identified residues critical for channel activation; we then generated binding modes for TRPA1-PIPC interactions using structural modeling, molecular docking, and mutational analysis. We show that PIPCs bind to a hydrophobic site located at the interface of the pore helix 1 (PH1) and S5 and S6 transmembrane segments. Interestingly, this binding site overlaps with that of known allosteric modulators, such as A-967079 and propofol. Similar binding sites, involving π-helix rearrangements on S6, have been recently reported for other TRP channels, suggesting an evolutionarily conserved mechanism. Finally, we show that for PIPC analogs, predictions from computational modeling are consistent with experimental structure-activity studies, thereby suggesting strategies for rational drug design.

19.
Artigo em Inglês | MEDLINE | ID: mdl-31814236

RESUMO

We report on a stabilizer of the interaction between 14-3-3ζ and the Estrogen Receptor alpha (ERα). ERα is a driver in the majority of breast cancers and 14-3-3 proteins are negative regulators of this nuclear receptor, making the stabilization of this protein-protein interaction (PPI) an interesting strategy. The stabilizer (1) consists of three symmetric peptidic arms containing an arginine mimetic, previously described as the GCP motif. 1 stabilizes the 14-3-3ζ/ERα interaction synergistically with the natural product Fusicoccin-A and was thus hypothesized to bind to a different site. This is supported by computational analysis of 1 binding to the binary complex of 14-3-3 and an ERα-derived phosphopeptide. Furthermore, 1 shows selectivity towards 14-3-3ζ/ERα interaction over other 14-3-3 client-derived phosphomotifs. These data provide a solid support of a new binding mode for a supramolecular 14-3-3ζ/ERα PPI stabilizer.

20.
Front Chem ; 7: 743, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31803713

RESUMO

Planar-chiral pillar[5]arenes bearing ß-D-galactose substituents on both rims have been successfully synthesized and effectively separated by silica gel chromatography with a high yield. The obtained (S p )- and (R p )-ß-D-galactose functionalized pillar[5]arenes [(S p-D )-GP5 and (R p-D )-GP5] exhibit the S p and R p planar chirality. Furthermore, (S p-D )-GP5 and (R p-D )-GP5 can not racemize according to dynamic 1H NMR and CD spectra. Notably, GP5 is able to capture a guest molecule (DNS-CPT) to form a host-guest supramolecular amphiphile, which can further self-assemble into chiral nanoparticles with the S p and R p planar chirality of (S p-D )-GP5 and (R p-D )-GP5 still being retained, suggesting GP5 could be as reliable chiral sources to transfer the S p and R p planar chirality.

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