Forecasting the energy demand and CO2 emissions of industrial sectors in China's Beijing-Tianjin-Hebei region under energy transition.

As the world's greatest energy consumer, China's energy consumption and transition have become a focus of attention. The most significant location for regional integration in the north of China is the Beijing-Tianjin-Hebei region, where the industrial sector dominates its energy consumption. Forecasting the energy demand and structure of industrial sectors in China's Beijing-Tianjin-Hebei region may help to promote the energy transition and CO2 emission mitigation. This study conducts a model based on the year 2020 using the Long-Range Energy Alternatives Planning System (LEAP) software and sets two scenarios (baseline scenario and emission peak scenario) to forecast the future energy demand and CO2 emissions of industrial sectors in China's Beijing-Tianjin-Hebei region until the year 2035. Moreover, the industrial sectors are classified into traditional high-energy-consuming industries, emerging manufacturing industries, daily-related light industries, and other industries. The forecasting results show that (1) The industrial energy demand of the entire Beijing-Tianjin-Hebei region will grow from 234 Mtce in 2020 to 317 Mtce in 2035, and the corresponding energy structure will shift from coal-based to electricity-based; (2) at the provincial level, all three provinces will experience an increase in industrial energy demand between 2020 and 2035, with Hebei experiencing the fastest average annual growth rate of 2.18% and the largest share of over 80%, and Beijing experiencing the highest average annual electrification rate of 70%; (3) at the industrial sector level, the electricity and natural gas will gradually replace other energy sources as the main energy source for industry. The most representative industrial sub-sector in Beijing, Tianjin, and Hebei provinces are all traditional high-energy-consuming industries, which will account for more than 90% of the total energy demand in both Tianjin and Hebei by 2035.

CCL28 Enhances HSV-2 gB-Specific Th1-Polarized Immune Responses against Lethal Vaginal Challenge in Mice.

Plasmid DNA (pDNA) represents a promising "genetic vaccine platform" capable of overcoming major histocompatibility complex barriers. We previously demonstrated that low-to-moderate doses of mucosae-associated epithelial chemokine (MEC or CCL28) as an immunomodulatory adjuvant can trigger effective and long-lasting systemic and mucosal HSV-2 gD-specific immune responses, whereas mice immunized with gD in combination with high-dose CCL28 showed toxicity and lost their immunoprotective effects after lethal HSV-2 challenge. The exact causes underlying high-dose, CCL28-induced lesions remain unknown. In an intramuscularly immunized mouse model, we investigated the immune-enhancement mechanisms of low-dose CCL28 as a molecular adjuvant combined with the relatively weak immunogen HSV-2 gB. Compared with the plasmid gB antigen group, we found that a low-dose of plasmid CCL28 (pCCL28) codelivered with pgB induced increased levels of gB-specific serum IgG and vaginal fluid IgA, serum neutralizing antibodies (NAb), Th1-polarized IgG2a, and cytokine IL-2 (>5-fold). Furthermore, low-dose pCCL28 codelivery with pgB enhanced CCL28/CCR10-axis responsive CCR10− plus CCR10+ B-cell (~1.2-fold) and DC pools (~4-fold) in the spleen, CCR10− plus CCR10+ T-cell pools (~2-fold) in mesenteric lymph nodes (MLNs), and the levels of IgA-ASCs in colorectal mucosal tissues, leading to an improved protective effect against a lethal dose of HSV-2 challenge. Findings in this study provide a basis for the development of CCL28-adjuvant vaccines against viral mucosal infections.

A Bright Monomeric Near-Infrared Fluorescent Protein with an Excitation Peak at 633 nm for Labeling Cellular Protein and Reporting Protein-Protein Interaction.

Bright monomeric near-infrared fluorescent proteins (NIR-FPs) are useful as markers for labeling proteins and cells and as sensors for reporting molecular activities in living cells and organisms. However, current monomeric NIR-FPs are dim under excitation with common 633/635/640 nm lasers, limiting their broad use in cellular/subcellular level imaging. Here, we report a bright monomeric NIR-FP with maximum excitation at 633 nm, named mIFP663, engineered from Xanthomonas campestris pv Campestris phytochrome (XccBphP). mIFP663 has high molecular brightness with a large extinction coefficient (86,600 M-1 cm-1) and a decent quantum yield (19.4%), and high cellular brightness that is 3-6 times greater than those of spectrally similar NIR-FPs in HEK293T cells in the presence of exogenous BV. Moreover, we demonstrate that mIFP663 is able to label critical cellular and viral proteins without perturbing subcellular localization and virus replication, respectively. Finally, with mIFP663, we engineer improved bimolecular fluorescence complementation (BiFC) and new bioluminescent resonance energy transfer (BRET) systems to detect protein-protein interactions in living cells.

FRET Imaging of Rho GTPase Activity with Red Fluorescent Protein-Based FRET Pairs.

With the development of fluorescent proteins (FPs) and advanced optical microscopy techniques, Förster or fluorescence resonance energy transfer (FRET) has become a powerful tool for real-time noninvasive visualization of a variety of biological processes, including kinase activities, with high spatiotemporal resolution in living cells and organisms. FRET can be detected in appropriately configured microscopes as changes in fluorescence intensity, lifetime, and anisotropy. Here, we describe the preparation of samples expressing FP-based FRET sensors for RhoA kinase, intensity- and lifetime-based FRET imaging, and postimaging data analysis.

Herpes Simplex Virus Type 2 Glycoprotein D Inhibits NF-κB Activation by Interacting with p65.

NF-κB plays a crucial role in regulating cell proliferation, inflammation, apoptosis, and immune responses. HSV type 2 (HSV-2) is one of the most predominant sexually transmitted pathogens worldwide, and its infection increases the risk of HIV type 1 (HIV-1) acquisition and transmission. HSV-2 glycoprotein D (gD), highly homologous to HSV-1 gD, is essential for viral adhesion, fusion, entry, and spread. It is known that HSV-1 gD can bind herpesvirus entry mediator (HVEM) to trigger NF-κB activation and thereby facilitate viral replication at the early stage of infection. In this study, we found that purified HSV-2 gD triggered NF-κB activation at the early stage of infection, whereas ectopic expression of HSV-2 gD significantly downregulated TNF-α-induced NF-κB activity as well as TNF-α-induced IL-6 and IL-8 expression. Mechanistically, HSV-2 gD inhibited NF-κB, but not IFN-regulatory factor 3 (IRF3), activation and suppressed NF-κB activation mediated by overexpression of TNFR-associated factor 2 (TRAF2), IκB kinase α (IKKα), IKKß, or p65. Coimmunoprecipitation and binding kinetic analyses demonstrated that HSV-2 gD directly bound to the NF-κB subunit p65 and abolished the nuclear translocation of p65 upon TNF-α stimulation. Mutational analyses further revealed that HSV-2 gD interacted with the region spanning aa 19-187 of p65. Findings in this study together demonstrate that HSV-2 gD interacts with p65 to regulate p65 subcellular localization and thereby prevents NF-κB-dependent gene expression, which may contribute to HSV-2 immune evasion and pathogenesis.

CCL19 and CCL28 Assist Herpes Simplex Virus 2 Glycoprotein D To Induce Protective Systemic Immunity against Genital Viral Challenge.

Potent systemic immunity is important for recalled mucosal immune responses, but in the defense against mucosal viral infections, it usually remains low at mucosal sites. Based on our previous findings that enhanced immune responses can be achieved by immunization with an immunogen in combination with a molecular adjuvant, here we designed chemokine-antigen (Ag) fusion constructs (CCL19- or CCL28-herpes simplex virus 2 glycoprotein D [HSV-2 gD]). After intramuscular (i.m.) immunization with different DNA vaccines in a prime and boost strategy, BALB/c mice were challenged with a lethal dose of HSV-2 through the genital tract. Ag-specific immune responses and chemokine receptor-specific lymphocytes were analyzed to determine the effects of CCL19 and CCL28 in strengthening humoral and cellular immunity. Both CCL19 and CCL28 were efficient in inducing long-lasting HSV-2 gD-specific systemic immunity. Compared to CCL19, less CCL28 was required to elicit HSV-2 gD-specific serum IgA responses, Th1- and Th2-like responses of immunoglobulin (Ig) subclasses and cytokines, and CCR3+ T cell enrichment (>8.5-fold) in spleens. These findings together demonstrate that CCL28 tends to assist an immunogen to induce more potently protective immunity than CCL19. This work provides information for the application potential of a promising vaccination strategy against mucosal infections caused by HSV-2 and other sexually transmitted viruses.IMPORTANCE An effective HSV-2 vaccine should induce antigen (Ag)-specific immune responses against viral mucosal infection. This study reveals that chemokine CCL19 or CCL28 enhanced HSV-2 glycoprotein D ectodomain (gD-306aa)-induced immune responses against vaginal virus challenge. In addition to eliciting robust humoral immune responses, the chemokine-Ag fusion construct also induced Th1- and Th2-like immune responses characterized by the secretion of multiple Ig subclasses and cytokines that were able to be recalled after HSV-2 challenge, while CCL28 appeared to be more effective than CCL19 in promoting gD-elicited immune responses as well as the migration of T cells to secondary lymph tissues. Of importance, both CCL19 and CCL28 significantly facilitated gD to induce protective mucosal immune responses in the genital tract. The above-described findings together highlight the potential of CCL19 or CCL28 in combination with gD as a vaccination strategy to control HSV-2 infection.

HSV-2 Infection of Human Genital Epithelial Cells Upregulates TLR9 Expression Through the SP1/JNK Signaling Pathway.

It is known that herpes simplex virus type 2 (HSV-2) triggers the activation of Toll-like receptor (TLR) 9 signaling pathway and the consequent production of antiviral cytokines in dendritic cells. However, the impact of HSV-2 infection on TLR9 expression and signaling in genital epithelial cells, the primary HSV-2 targets, has yet to be determined. In the current study, by using both human genital epithelial cell lines and primary genital epithelial cells as models, we found that HSV-2 infection enhances TLR9 expression at both mRNA and protein levels. Such enhancement is virus replication-dependent and CpG-independent, while the HSV-2-mediated upregulation of TLR9 does not activate TLR9 signaling pathway. Mechanistically, a SP1 binding site on TLR9 promoter appears to be essential for HSV-2-induced TLR9 transactivation. Upon HSV-2 infection, SP1 translocates from the cytoplasm to the nucleus, and consequently binds to TLR9 promoter. By using specific inhibitors, the JNK signaling pathway is shown to be involved in the HSV-2-induced TLR9 transactivation, while HSV-2 infection increases the phosphorylation but not the total level of JNK. In agreement, antagonism of JNK signaling pathway inhibits the HSV-2-induced SP1 nuclear translocation. Taken together, our study demonstrates that HSV-2 infection of human genital epithelial cells promotes TLR9 expression through SP1/JNK signaling pathway. Findings in this study provide insights into HSV-2-host interactions and potential targets for immune intervention.

Herpes Simplex Virus Type 2 Immediate Early Protein ICP27 Inhibits IFN-ß Production in Mucosal Epithelial Cells by Antagonizing IRF3 Activation.

Herpes simplex virus type 2 (HSV-2) is the main cause of genital herpes and infections are common in the lower genital tract. Although neuronal and immune cells can be infected, epithelial cells, and keratinocytes are the primary HSV-2 target cells. HSV-2 establishes latency by evading the host immune system and its infection can also increase the risk of HIV-1 sexual transmission. Our pervious study found that HSV-2 immediate early protein ICP22, inhibited IFN-ß production by interfering with the IRF3 pathway. However, ICP22-null HSV-2 did not completely lose the capability of suppressing IFN-ß induction, suggesting the involvement of other viral components in the process. In this study, by using an ex vivo cervical explant model, we first demonstrated that HSV-2 can indeed inhibit IFN-ß induction in human mucosal tissues. We further identified HSV-2 immediate early protein ICP27 as a potent IFN-ß antagonist. ICP27 significantly suppresses the Sendai virus or polyinosinic-polycytidylic acid-induced IFN-ß production in human mucosal epithelial cells, showing that ICP27 inhibits the IFN-ß promoter activation, and IFN-ß production at both mRNA and protein levels. Additional studies revealed that ICP27 directly associates with IRF3 and inhibits its phosphorylation and nuclear translocation, resulting in the inhibition of IFN-ß induction. Our findings provide insights into the molecular mechanism underlying HSV-2 mucosal immune evasion, and information for the design of HSV-2 mucosal vaccines.

HSV-2 glycoprotein J promotes viral protein expression and virus spread.

HSV-2 spread is predominantly dependent on cell-to-cell contact. However, the underlying mechanisms remain to be determined. Here we demonstrate that HSV-2 gJ, which was previously assigned no specific function, promotes HSV-2 cell-to-cell spread and syncytia formation. In the context of viral infection, knockout or knockdown of gJ impairs HSV-2 cell-to-cell spread among epithelial cells or from epithelial cells to neuronal cells, which leads to decreased virus production, whereas ectopic expression of gJ enhances virus production. Mechanistically, gJ increases the expression levels of HSV-2 proteins, and also enhances viral protein expression and replication of heterologous viruses like HIV-1 and JEV, suggesting that HSV-2 gJ likely functions as a regulator of viral protein expression and virus production. Findings in this study provide a basis for further understanding the role of gJ in HSV-2 replication.

Penton base induces better protective immune responses than fiber and hexon as a subunit vaccine candidate against adenoviruses.

Human adenoviruses (AdVs) have been extensively studied as vectors for gene therapy and vaccination. However, little attention has been paid to AdV vaccine development and treatment. Currently, there is a lack of information concerning the immunogenicity of AdV major capsid proteins. Here, using AdV7 as a model, we compared the immunogenicity and protection efficacy of its three major capsid proteins in DNA forms, pFiber, pHexon and pPenton, on a mouse model. Quantification of antigen-specific antibodies showed that pHexon induced highest IgG in sera while pPenton induced highest IgA in respiratory mucosae. A neutralization assay revealed that pPenton elicited the highest neutralizing activity against the homologous AdV7 in both sera and bronchoalveolar lavages (BALs). In addition, sera and BALs from mice immunized with either of the three constructs had cross-neutralizing activities against the heterologous AdV3. Furthermore, pHexon and pPenton induced Th1/2- and Th1/17-biased cellular responses, respectively, with pFiber being the weakest in the induction of cellular responses. Virus challenge assay showed that, pPenton group had the fastest virus clearance rate, followed by pFiber and pHexon groups. Likewise, the inflammation in the lung was well controlled in pPenton group against virus challenge. Taken together, our data demonstrate that penton base is better than fiber and hexon as a vaccine candidate against AdVs. Our findings provide important information for the development of subunit vaccines against AdVs.

Herpes Simplex Virus Type 2 Infection-Induced Expression of CXCR3 Ligands Promotes CD4+ T Cell Migration and Is Regulated by the Viral Immediate-Early Protein ICP4.

HSV-2 infection-induced CXCR3 ligands are important for the recruitment of virus-specific CD8+ T cells, but their impact on CD4+ T cell trafficking remains to be further determined. Given that recruitment of CD4+ T cells to infection areas may be one of the mechanisms that account for HSV-2 infection-mediated enhancement of HIV-1 sexual transmission, here we investigated the functionality of HSV-2 infection-induced CXCR3 ligands CXCL9, CXCL10, and CXCL11 in vivo and in vitro, and determined the viral components responsive for such induction and the underlying mechanisms. We first found that the expression of CXCR3 ligands CXCL9, CXCL10, and CXCL11 was increased in mice following vaginal challenge with HSV-2, while CXCL9 played a predominant role in the recruitment of CD4+ T cells to the vaginal foci of infected mice. HSV-2 infection also induced the production of CXCL9, CXCL10, and CXCL11 in human cervical epithelial cells. Of note, although HSV-2 induced the expression of all the three CXCR3 ligands, the induced CXCL9 appeared to play a predominant role in promoting CD4+ T cell migration, reflecting that the concentrations of CXCL10 and CXCL11 required for CD4+ T cell migration are higher than that of CXCL9. We further revealed that, ICP4, an immediate-early protein of HSV-2, is crucial in promoting CXCR3 ligand expression through the activation of p38 MAPK pathway. Mechanistically, ICP4 binds to corresponding promoters of CXCR3 ligands via interacting with the TATA binding protein (TBP), resulting in the transcriptional activation of the corresponding promoters. Taken together, our study highlights HSV-2 ICP4 as a vital viral protein in promoting CXCR3 ligand expression and CXCL9 as the key induced chemokine in mediating CD4+ T cell migration. Findings in this study have shed light on HSV-2 induced leukocyte recruitment which may be important for understanding HSV-2 infection-enhanced HIV-1 sexual transmission and the development of intervention strategies.

Japanese encephalitis virus counteracts BST2 restriction via its envelope protein E.

It has been well documented that BST2 restricts the release of enveloped viruses by cross-linking newly produced virions to the cell membrane. However, it is less clear whether and how BST2 inhibits the release of enveloped viruses which bud via the secretory pathway. Here, we demonstrated that BST2 restricts the release of Japanese encephalitis virus (JEV) whose budding occurs at the ER-Golgi intermediate compartment, and in turn, JEV infection downregulates BST2 expression. We further found that the JEV envelope protein E, but not other viral components, significantly downregulates BST2 with the viral protein M playing an auxiliary role in the process. Envelope protein E-mediated BST2 downregulation appears to undergo lysosomal degradation pathway. Additional study revealed that the transmembrane domain and the coiled-coil domain (CC) of BST2 are the target domains of viral protein E and that the N- and C-terminal membrane anchors and the CC domain of BST2 are essential for blocking JEV release. Our results together indicate that the release of enveloped viruses whose budding take place in an intracellular compartment can be restricted by BST2.

Immunization with HSV-2 gB-CCL19 Fusion Constructs Protects Mice against Lethal Vaginal Challenge.

There is a lack of an HSV-2 vaccine, in part as the result of various factors that limit robust and long-term memory immune responses at the mucosal portals of viral entry. We previously demonstrated that chemokine CCL19 augmented mucosal and systemic immune responses to HIV-1 envelope glycoprotein. Whether such enhanced immunity can protect animals against virus infection remains to be addressed. We hypothesized that using CCL19 in a fusion form to direct an immunogen to responsive immunocytes might have an advantage over CCL19 being used in combination with an immunogen. We designed two fusion constructs, plasmid (p)gBIZCCL19 and pCCL19IZgB, by fusing CCL19 to the C- or N-terminal end of the extracellular HSV-2 glycoprotein B (gB) with a linker containing two (Gly4Ser)2 repeats and a GCN4-based isoleucine zipper motif for self-oligomerization. Following immunization in mice, pgBIZCCL19 and pCCL19IZgB induced strong gB-specific IgG and IgA in sera and vaginal fluids. The enhanced systemic and mucosal Abs showed increased neutralizing activity against HSV-2 in vitro. Measurement of gB-specific cytokines demonstrated that gB-CCL19 fusion constructs induced balanced Th1 and Th2 cellular immune responses. Moreover, mice vaccinated with fusion constructs were well protected from intravaginal lethal challenge with HSV-2. Compared with pgB and pCCL19 coimmunization, fusion constructs increased mucosal surface IgA(+) cells, as well as CCL19-responsive immunocytes in spleen and mesenteric lymph nodes. Our findings indicate that enhanced humoral and cellular immune responses can be achieved by immunization with an immunogen fused to a chemokine, providing information for the design of vaccines against mucosal infection by HSV-2 and other sexually transmitted viruses.

Functional characterization of a mycothiol peroxidase in Corynebacterium glutamicum that uses both mycoredoxin and thioredoxin reducing systems in the response to oxidative stress.

Previous studies have identified a putative mycothiol peroxidase (MPx) in Corynebacterium glutamicum that shared high sequence similarity to sulfur-containing Gpx (glutathione peroxidase; CysGPx). In the present study, we investigated the MPx function by examining its potential peroxidase activity using different proton donors. The MPx degrades hydrogen peroxide and alkyl hydroperoxides in the presence of either the thioredoxin/Trx reductase (Trx/TrxR) or the mycoredoxin 1/mycothione reductase/mycothiol (Mrx1/Mtr/MSH) regeneration system. Mrx1 and Trx employ different mechanisms in reducing MPx. For the Mrx1 system, the catalytic cycle of MPx involves mycothiolation/demycothiolation on the Cys(36) sulfenic acid via the monothiol reaction mechanism. For the Trx system, the catalytic cycle of MPx involves formation of an intramolecular disulfide bond between Cys(36) and Cys(79) that is pivotal to the interaction with Trx. Both the Mrx1 pathway and the Trx pathway are operative in reducing MPx under stress conditions. Expression of mpx markedly enhanced the resistance to various peroxides and decreased protein carbonylation and intracellular reactive oxygen species (ROS) accumulation. The expression of mpx was directly activated by the stress-responsive extracytoplasmic function-σ (ECF-σ) factor [SigH]. Based on these findings, we propose that the C. glutamicum MPx represents a new type of GPx that uses both mycoredoxin and Trx systems for oxidative stress response.

Inhibition of HIV-1 infection of primary CD4+ T-cells by gene editing of CCR5 using adenovirus-delivered CRISPR/Cas9.

CCR5 serves as an essential coreceptor for human immunodeficiency virus type 1 (HIV-1) entry, and individuals with a CCR5(Δ32) variant appear to be healthy, making CCR5 an attractive target for control of HIV-1 infection. The CRISPR/Cas9, which functions as a naturally existing adaptive immune system in prokaryotes, has been recently harnessed as a novel nuclease system for genome editing in mammalian cells. Although CRISPR/Cas9 can be readily delivered into cell lines, due to the large size of the Cas9 protein, efficient delivery of CCR5-targeting CRISPR/Cas9 components into primary cells, including CD4(+) T-cells, the primary target for HIV-1 infection in vivo, remains a challenge. In the current study, following design of a panel of top-ranked single-guided RNAs (sgRNAs) targeting the ORF of CCR5, we demonstrate that CRISPR/Cas9 can efficiently mediate the editing of the CCR5 locus in cell lines, resulting in the knockout of CCR5 expression on the cell surface. Next-generation sequencing revealed that various mutations were introduced around the predicted cleavage site of CCR5. For each of the three most effective sgRNAs that we analysed, no significant off-target effects were detected at the 15 top-scoring potential sites. More importantly, by constructing chimeric Ad5F35 adenoviruses carrying CRISPR/Cas9 components, we efficiently transduced primary CD4(+) T-lymphocytes and disrupted CCR5 expression, and the positively transduced cells were conferred with HIV-1 resistance. To our knowledge, this is the first study establishing HIV-1 resistance in primary CD4(+) T-cells utilizing adenovirus-delivered CRISPR/Cas9.

HSV-2 immediate-early protein US1 inhibits IFN-ß production by suppressing association of IRF-3 with IFN-ß promoter.

HSV-2 is the major cause of genital herpes, and its infection increases the risk of HIV-1 acquisition and transmission. After initial infection, HSV-2 can establish latency within the nervous system and thus maintains lifelong infection in humans. It has been suggested that HSV-2 can inhibit type I IFN signaling, but the underlying mechanism has yet to be determined. In this study, we demonstrate that productive HSV-2 infection suppresses Sendai virus (SeV) or polyinosinic-polycytidylic acid-induced IFN-ß production. We further reveal that US1, an immediate-early protein of HSV-2, contributes to such suppression, showing that US1 inhibits IFN-ß promoter activity and IFN-ß production at both mRNA and protein levels, whereas US1 knockout significantly impairs such capability in the context of HSV-2 infection. US1 directly interacts with DNA binding domain of IRF-3, and such interaction suppresses the association of nuclear IRF-3 with the IRF-3 responsive domain of IFN-ß promoter, resulting in the suppression of IFN-ß promoter activation. Additional studies demonstrate that the 217-414 aa domain of US1 is critical for the suppression of IFN-ß production. Our results indicate that HSV-2 US1 downmodulates IFN-ß production by suppressing the association of IRF-3 with the IRF-3 responsive domain of IFN-ß promoter. Our findings highlight the significance of HSV-2 US1 in inhibiting IFN-ß production and provide insights into the molecular mechanism by which HSV-2 evades the host innate immunity, representing an unconventional strategy exploited by a dsDNA virus to interrupt type I IFN signaling pathway.