Dengue virus (DENV) infects millions of people worldwide and is a major public health problem. DENV nonstructural protein 1 (NS1) is a conserved glycoprotein that associates with membranes and is also secreted into the plasma in DENV-infected patients. The present study describes a novel mechanism by which NS1 inhibits the terminal complement pathway. We first identified the terminal complement regulator vitronectin (VN) as a novel DENV2 NS1 binding partner by using a yeast two-hybrid system. This interaction was further assessed by enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR) assay. The NS1-VN complex was also detected in plasmas from DENV-infected patients, suggesting that this interaction occurs during DENV infection. We also demonstrated that the DENV2 NS1 protein, either by itself or by interacting with VN, hinders the formation of the membrane attack complex (MAC) and C9 polymerization. Finally, we showed that DENV2, West Nile virus (WNV), and Zika virus (ZIKV) NS1 proteins produced in mammalian cells inhibited C9 polymerization. Taken together, our results points to a role for NS1 as a terminal pathway inhibitor of the complement system. IMPORTANCE Dengue is the most important arthropod-borne viral disease nowadays and is caused by dengue virus (DENV).The flavivirus NS1 glycoprotein has been characterized functionally as a complement evasion protein that can attenuate the activation of the classical, lectin, and alternative pathways. The present study describes a novel mechanism by which DENV NS1 inhibits the terminal complement pathway. We identified the terminal complement regulator vitronectin (VN) as a novel DENV NS1 binding partner, and the NS1-VN complex was detected in plasmas from DENV-infected patients, suggesting that this interaction occurs during DENV infection. We also demonstrated that the NS1-VN complex inhibited membrane attack complex (MAC) formation, thus interfering with the complement terminal pathway. Interestingly, NS1 itself also inhibited MAC activity, suggesting a direct role of this protein in the inhibition process. Our findings imply a role for NS1 as a terminal pathway inhibitor of the complement system. D engue constitutes a major public health problem in tropical and subtropical countries. According to current estimates, at least 390 million cases of dengue occur annually, of which approximately 100 million are symptomatic (1). The infection is caused by dengue virus (DENV), a member of the Flaviviridae family that cocirculates in nature as four distinct antigenic serotypes (DENV1 to -4). DENV infection in humans is generally asymptomatic, but symptomatic cases can vary from a mild and self-limited fever to a potentially fatal hemorrhagic syndrome (2). The DENV genome is composed of a single positive-sense RNA that encodes a single viral polyprotein that is further processed by viral and host proteases into three structural proteins (C, prM/M, and E) and seven nonstructural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) (3).NS1 ...

Dengue is one of the main public health concerns worldwide. Recent estimates indicate that over 390 million people are infected annually with the dengue virus (DENV), resulting in thousands of deaths. Among the DENV nonstructural proteins, the NS1 protein is the only one whose function during replication is still unknown. NS1 is a 46-to 55-kDa glycoprotein commonly found as both a membrane-associated homodimer and a soluble hexameric barrel-shaped lipoprotein. Despite its role in the pathogenic process, NS1 is essential for proper RNA accumulation and virus production. In the present study, we identified that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) interacts with intracellular NS1. Molecular docking revealed that this interaction occurs through the hydrophobic protrusion of NS1 and the hydrophobic residues located at the opposite side of the catalytic site. Moreover, addition of purified recombinant NS1 enhanced the glycolytic activity of GAPDH in vitro. Interestingly, we observed that DENV infection promoted the relocalization of GAPDH to the perinuclear region, where NS1 is commonly found. Both DENV infection and expression of NS1 itself resulted in increased GAPDH activity. Our findings indicate that the NS1 protein acts to increase glycolytic flux and, consequently, energy production, which is consistent with the recent finding that DENV induces and requires glycolysis for proper replication. This is the first report to propose that NS1 is an important modulator of cellular energy metabolism. The data presented here provide new insights that may be useful for further drug design and the development of alternative antiviral therapies against DENV. IMPORTANCE Dengue represents a serious public health problem worldwide and is caused by infection with dengue virus (DENV).Estimates indicate that half of the global population is at risk of infection, with almost 400 million cases occurring per year. The NS1 glycoprotein is found in both the intracellular and the extracellular milieus. Despite the fact that NS1 has been commonly associated with DENV pathogenesis, it plays a pivotal but unknown role in the replication process. In an effort to understand the role of intracellular NS1, we demonstrate that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) interacts with NS1. Our results indicate that NS1 increases the glycolytic activity of GAPDH in vitro. Interestingly, the GAPDH activity was increased during DENV infection, and NS1 expression alone was sufficient to enhance intracellular GAPDH activity in BHK-21 cells. Overall, our findings suggest that NS1 is an important modulator of cellular energy metabolism by increasing glycolytic flux. Dengue is one of the major health problems in tropical regions. It is estimated that over 390 million people are infected annually with one of the four dengue virus (DENV) serotypes (1). The absence of both an effective tetravalent vaccine and therapeutic agents worsens the impact of the dengue burden. DENV, the most threatening member of the Flaviviridae family,...

Dengue virus (DENV) represents a major threat to public health worldwide. Early DENV diagnosis should not only detect the infection but also identify patients with a higher likelihood to develop severe cases. Previous studies have suggested the potential for NS1 to serve as a viral marker for dengue severity. However, further studies using different sera panels are required to confirm this hypothesis. In this context, we developed a lab-based ELISA to detect and quantitate NS1 protein from the four DENV serotypes and from primary and secondary cases. This approach was used to calculate the circulating NS1 concentration in positive samples. We also tested the NS1 positivity of DENV-positive samples according to the Platelia Dengue NS1 Ag assay. A total of 128 samples were positive for DENV infection and were classified according to the WHO guidelines. The overall NS1 positivity was 68% according to the Platelia assay, whereas all samples were NS1-positive when analyzed with our lab-based ELISA. Fifty-four samples were positive by PCR, revealing a co-circulation of DENV1 and DENV4, and the NS1 positivity for DENV4 samples was lower than that for DENV1. The circulating NS1 concentration ranged from 7 to 284 ng/mL. Our results support previous data indicating the low efficiency of the Platelia assay to detect DENV4 infection. Moreover, this work is the first to analyze NS1 antigenemia using retrospective samples from a Brazilian outbreak.

Dengue is a mosquito-borne infectious disease highly endemic to tropical and subtropical countries. Symptomatic patients can rapidly progress to severe conditions of hemorrhage, plasma extravasation and hypovolemic shock that leads to death. Severe dengue patients’ blood tests typically report low levels of high-density lipoprotein (HDL), that is responsible for reverse cholesterol transport (RCT) and regulation of lipid composition on peripheral tissues. It is well known that Dengue virus (DENV) depends on membrane cholesterol rafts to infect and replicate in mammalian cells. Here we describe the interaction of the DENV nonstructural protein 1 (NS1) with Apolipoprotein A1 (ApoA1), which is the major protein component of HDL. NS1 is secreted by infected cells and can be found circulating in the serum of patients since the onset of the symptoms. NS1 concentration in plasma is related to dengue severity, attributed to immune evasion and acute inflammatory response. Our data show that the DENV NS1 protein induces the increase of lipid rafts in non-infected cell membrane and enhances further DENV infection. We also show that ApoA1-mediated lipid raft depletion inhibits DENV attachment to cell surface. In addition, ApoA1 was able to neutralize NS1-induced cell activation and to prevent NS1-mediated enhancement of DENV infection. Furthermore, we demonstrate that the ApoA1 mimetic peptide 4F is also capable of mediating lipid raft depletion to control DENV infection. Taken together, our results suggest the potential of RCT-based therapies for dengue treatment. These results should motivate studies to assess the importance of RCT in dengue infection in vivo. IMPORTANCE Dengue virus (DENV) is one of the most relevant mosquito-transmitted viruses worldwide, infecting more the 390 million people every year and leading to more than 20 thousand deaths. Despite a DENV vaccine has already been approved, its potential side-effects have hampered its use in large scale immunization. Therefore, new treatment options are urgent either to prevent disease worsening or improve current clinical management of severe cases. In the present study, we describe a new interaction of the NS1 protein, one of the major viral components, with a key component of high-density lipoprotein, the Apolipoprotein A1 (ApoA1). This interaction seems to alter membrane susceptibility to virus infection and modulates the mechanisms triggered by DENV to evade immune response. We also propose the use of a mimetic peptide named 4F, originally developed for atherosclerosis, as a potential therapy for relieving DENV symptoms.

In chronic schistosomiasis, liver fibrosis is linked to portal hypertension, which is a condition associated with high mortality and morbidity. High mobility group box 1 (HMGB1) was originally described as a nuclear protein that functions as a structural co-factor in transcriptional regulation. However, HMGB1 can also be secreted into the extracellular milieu under appropriate signal stimulation. Extracellular HMGB1 acts as a multifunctional cytokine that contributes to infection, injury, inflammation, and immune responses by binding to specific cell-surface receptors. HMGB1 is involved in fibrotic diseases. From a clinical perspective, HMGB1 inhibition may represent a promising therapeutic approach for treating tissue fibrosis. In this study, we demonstrate elevated levels of HMGB1 in the sera in experimental mice or in patients with schistosomiasis. Using immunohistochemistry, we demonstrated that HMGB1 trafficking in the hepatocytes of mice suffering from acute schistosomiasis was inhibited by Glycyrrhizin, a well-known HMGB1 direct inhibitor, as well as by DIC, a novel and potential anti-HMGB1 compound. HMGB1 inhibition led to significant downregulation of IL-6, IL4, IL-5, IL-13, IL-17A, which are involved in the exacerbation of the immune response and liver fibrogenesis. Importantly, infected mice that were treated with DIC or GZR to inhibit HMGB1 pro-inflammatory activity showed a significant increase in survival and a reduction of over 50% in the area of liver fibrosis. Taken together, our findings indicate that HMGB1 is a key mediator of schistosomotic granuloma formation and liver fibrosis and may represent an outstanding target for the treatment of schistosomiasis.