Endosomes are characterized by the presence of various phosphoinositides that are essential for defining the membrane properties. However, the interplay between endosomal phosphoinositides metabolism and innate immunity is yet to be fully understood. Here, our findings highlight the evolutionary continuity of RAB-10/Rab10's involvement in regulating innate immunity. Upon infection of Caenorhabditis elegans with Pseudomonas aeruginosa, an increase in RAB-10 activity was observed in the intestine. Conversely, when RAB-10 was absent, the intestinal diacylglycerols (DAGs) decreased, and the animal's response to the pathogen was impaired. Further research revealed that UNC-16/JIP3 acts as an RAB-10 effector, facilitating the recruitment of phospholipase EGL-8 to endosomes. This leads to a decrease in endosomal phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and an elevation of DAGs, as well as the activation of the PMK-1/p38 MAPK innate immune pathway. It is noteworthy that the dimerization of UNC-16 is a prerequisite for its interaction with RAB-10(GTP) and the recruitment of EGL-8. Moreover, we ascertained that the rise in RAB-10 activity, due to infection, was attributed to the augmented expression of LET-413/Erbin, and the nuclear receptor NHR-25/NR5A1/2 was determined to be indispensable for this increase. Hence, this study illuminates the significance of endosomal PI(4,5)P2 catabolism in boosting innate immunity and outlines an NHR-25-mediated mechanism for pathogen detection in intestinal epithelia.
Animals ; Caenorhabditis elegans/genetics* ; Endosomes/immunology* ; Caenorhabditis elegans Proteins/immunology* ; Phosphatidylinositol 4,5-Diphosphate/immunology* ; Immunity, Innate ; Pseudomonas aeruginosa/immunology* ; rab GTP-Binding Proteins/genetics* ; Diglycerides/metabolism*

To develop a safe and effective new generation vaccine for IRKV-THChina12 prevention, we constructed a non-replicative recombinant human adenovirus carrying the IRKV-THChina12 G gene, named as rAd5-IRKV-G. The IRKV-THChina12 G protein expressed by the recombinant human adenovirus in 293AD cells was detected by western blot and indirect immunofluorescence test. To evaluate the immunogenicity of the recombinant, mice were immunized with rAd5-IRKV-G by intramuscular (i. m.) or intraperitoneal (i. p.) route and with non-exogenous gene expressing wild type adenovirus wt-rAd5 as a control. Results showed that the rAd5-IRKV-G could induce continuous and statistically significant (P ≤ 0.05) anti-IRKV neutralizing antibody (NA) production in immunized mice by i. m. or i. p. route. In particular, no significant difference (P > 0.05) of the NA titers between the two administration routes were observed, that provides an alternative choice for animal immunization method in the future application.
Adenoviruses, Human ; genetics ; physiology ; Animals ; Antibodies, Neutralizing ; immunology ; Antibodies, Viral ; immunology ; GTP-Binding Proteins ; genetics ; immunology ; Gene Expression ; Genetic Vectors ; genetics ; physiology ; Humans ; Immunization ; Lyssavirus ; enzymology ; genetics ; immunology ; Mice ; Rhabdoviridae Infections ; immunology ; virology ; Viral Proteins ; genetics ; immunology ; Virus Replication

OBJECTIVETo review the mechanisms by which HIV evades different components of the host immune system.

DATA SOURCESThis review is based on data obtained from published articles from 1991 to 2012. To perform the PubMed literature search, the following key words were input: HIV and immune evasion.

STUDY SELECTIONArticles containing information related to HIV immune evasion were selected.

RESULTSAlthough HIV is able to induce vigorous antiviral immune responses, viral replication cannot be fully controlled, and neither pre-existing infected cells nor latent HIV infection can be completely eradicated. Like many other enveloped viruses, HIV can escape recognition by the innate and adaptive immune systems. Recent findings have demonstrated that HIV can also successfully evade host restriction factors, the components of intrinsic immune system, such as APOBEC3G (apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like 3G), TRIM5α (tripartite motif 5-α), tetherin, and SAMHD1 (SAM-domain HD-domain containing protein).

CONCLUSIONSHIV immune evasion plays an important role in HIV pathogenesis. Fully understanding the tactics deployed by HIV to evade various components of the host immune systems will allow for the development of novel strategies aimed toward the prevention and cure of HIV/AIDS.

APOBEC-3G Deaminase ; Adaptive Immunity ; Antibodies, Neutralizing ; immunology ; Antigens, CD ; physiology ; Carrier Proteins ; physiology ; Complement System Proteins ; immunology ; Cytidine Deaminase ; physiology ; GPI-Linked Proteins ; physiology ; HIV-1 ; immunology ; Humans ; Immune Evasion ; Killer Cells, Natural ; immunology ; Monomeric GTP-Binding Proteins ; physiology ; SAM Domain and HD Domain-Containing Protein 1

OBJECTIVETo investigate the possible influence of HBV and its antigens on the expressions of JAK-STAT signal transduction pathway molecules and the antiviral proteins of IFN alpha.

METHODSThe HepG2 cells were transfected with pSM2, pHBS2-S and pHBc-EGFP plasmids which express HBV whole particles or S-antigen, Pre-S antigen and core antigens. The infectious supernatant from HepG2.2.15 cells and the pured HBV proteins which contained the S, Pre-S antigens were used to treat the HepG2 cells. Northern blot and RT-PCR were applied to analyse the expressions of the antiviral proteins MxA, 2' -5' OAS, 9-27 and the JAK-STAT signal transduction pathway molecules STAT1 in HepG2 cells responded to the IFN alpha treatment.

RESULTSThe HepG2 cells transfected with pSM2, pHBS2-S and pHBc-EGFP plasmids could express whole HBV particles and HBsAg, Pre-S antigen and HBcAg. The quantitation of expressed HBV particles and antigens increased significantly during the course of transfection. Northern blot hybridization analysis indicated that the HepG2 cells expressed IFN alpha antiviral proteins MxA, 2' -5' OAS and 9-27. When transfected with pHBV-dimer, pHBS2-S, pHBc-EGFP plasmids, the IFN/A antiviral proteins MxA, 2' -5' OAS and 9-27 in transfected cells were reduced greatly as compared to the un-transfected HepG2 cells, and the expressed antiviral proteins decreased sharply with the development of transfection time. Furthermore, the expression of IFN alpha JAK-STAT signal transduction pathway molecule STAT1 was also inhibited with the expression of HBV particles and HBV antigens in transfected HepG2 cells.

CONCLUSIONSThe HBV and its antigens influence the expressions of IFN alpha JAK-STAT signal transduction pathway molecules and antiviral proteins in the hepatocellular models in vitro. It is indicated that HBV might possess the activity to antagonise or counteract the IFN alpha antiviral action.

2',5'-Oligoadenylate Synthetase ; metabolism ; GTP-Binding Proteins ; metabolism ; Hep G2 Cells ; Hepatitis B Antigens ; immunology ; Hepatitis B virus ; immunology ; Humans ; Interferon-alpha ; metabolism ; Myxovirus Resistance Proteins ; RNA-Binding Proteins ; metabolism ; STAT1 Transcription Factor ; metabolism ; Signal Transduction ; Transfection

Previously, we reported that CD40-induced production of reactive oxygen species (ROS) by NADPH oxidase requires the TNF receptor-associated factor (TRAF) 3, as well as the activities of phosphatidylinositol 3-kinase (PI3K) and Rac1. Here we investigated the possible mechanisms of the production of ROS after CD40 ligation in B cells. We describe an alternative ROS production pathway that is triggered by CD40 ligation, involves 5-lipoxygenase (5-LO), and results in activation of p38 MAPK. Our studies in Raji human B lymphomas revealed that CD40-induced ROS production by 5-LO also requires the activities of PI3K and Rac1. In contrast to the NADPH oxidase pathway, however, TRAF molecules are not required for the CD40-induced ROS production by 5-LO. The association of CD40 with 5-LO is dependent on CD40 ligation in Raji B cells, and co-immunoprecipitation experiments using epitope-tagged proteins transiently expressed in human embryonic kidney 293T cells revealed the role of the regulatory subunit of PI3K, p85, in this association. Collectively, these data suggest a separate pathway for the CD40-induced ROS production in B cells and demonstrate that this pathway requires 5-LO via direct association of p85 with both CD40 and 5-LO.
Antigens, CD40/*metabolism ; Arachidonate 5-Lipoxygenase/*metabolism ; B-Lymphocytes/*enzymology/immunology ; CD40 Ligand/metabolism ; Cell Line, Tumor ; *Enzyme Activation ; HEK293 Cells ; Humans ; Phosphatidylinositol 3-Kinases/metabolism ; Protein Binding ; *Reactive Oxygen Species/metabolism ; Signal Transduction ; p38 Mitogen-Activated Protein Kinases/*metabolism ; rac GTP-Binding Proteins/metabolism

OBJECTIVETo study the human myxovirus resistant protein A (MxA), a specifically induced peptide by interferon I, and to use its level as a diagnostic criterion for viral infections.

METHODSAnti-MxA antisera from immunized mice were prepared with the expressed MxA protein of pET32a-MxA in E. coli BL-21(DE3). To confirm the antiserum activity and specificity, the expression product of BL21, wild type MxA pEGFP-C1-wMxA and site-directed mutant MxA pEGFP-C1-mMxA(N589S) stably transfected 3T3 cells and induced A549 cells were detected by Western blot with the antisera using non-MxA transfected or non-IFN-beta induced cells, intact A549, NIH 3T3 cells transfected with pEGFP-C1 and pET32a (+)-transformed BL-21 as controls.

RESULTSThe antisera had specific positive immunoreactivity to the NIH3T3 cells transformed with pEGFP-C1-wMxA and pEGFP-C1-mMxA, INF-beta induced A549 cells and BL21 proteins expressed with pET32a (+)-MxA. The hybridization signals from IFN-beta induced A549 cells depended on the IFN-beta inducing concentrations. Meanwhile, immunohistochemical assay showed that NIH 3T3 cells with pEGFP-C1-wMxA and pEGFP-C1-mMxA had > 98% of positive cells at 1:50 dilution of the serum and A549 cells induced by 20 ng/mL IFN-beta for 48 h showed 95% positive cells. pEGFP-C1-transfected NIH 3T3 cells were all negative.

CONCLUSIONAnti-sera are highly specific to diversified MxAs. The antibody is detectable by Western blot, immunocytochemistry and immunofluorescence assay.

Animals ; Antibody Specificity ; Cell Line, Tumor ; GTP-Binding Proteins ; genetics ; immunology ; metabolism ; Gene Expression Regulation ; Humans ; Mice ; Myxovirus Resistance Proteins ; NIH 3T3 Cells ; Species Specificity

Intrinsic cellular defenses are non-specific antiviral activities by recognizing pathogen-associated molecular patterns (PAMPs). Toll-like receptors (TLRs), one of the pathogen recognize receptor (PRR), sense various microbial ligands. Especially, TLR2, TLR3, TLR4, TLR7, TLR8 and TLR9 recognize viral ligands such as glycoprotein, single- or double-stranded RNA and CpG nucleotides. The binding of viral ligands to TLRs transmits its signal to Toll/interleukin-1 receptor (TIR) to activate transcription factors via signal transduction pathway. Through activation of transcription factors, such as interferon regulatory factor-3, 5, and 7 (IRF-3, 5, 7) or nuclear factor-kappaB (NF-kappaB), type I interferons are induced, and antiviral proteins such as myxovirus-resistance protein (Mx) GTPase, RNA-dependent Protein Kinase (PKR), ribonuclease L (RNase L), Oligo-adenylate Synthetase (OAS) and Interferon Stimulated Gene (ISG) are further expressed. These antiviral proteins play an important role of antiviral resistancy against several viral pathogens in infected cells and further activate innate immune responses.
Animals ; GTP-Binding Proteins/metabolism ; Humans ; Interferon Regulatory Factors/metabolism ; Interferon Type I/*metabolism/physiology ; Models, Biological ; NF-kappa B/metabolism ; Toll-Like Receptors/metabolism ; Virus Diseases/*immunology/*metabolism/virology ; eIF-2 Kinase/metabolism

The aim of this study was to investigate the role of RhoA/mDia1 pathway in the process of thrombin-induced platelet aggregation and regulatory effect of PI3K inhibitor on this process. The human platelets were isolated from peripheral blood, the activation of RhoA, Rac1 and Cdc42 in the platelet aggregation was detected by GST pull-down assay and immune co-precipitation, the interaction of RhoA, Rac1 and Cdc42 with mDia1 and the formation of complex in the process of platelet aggregation were determined by immune coprecipitation, and the effect of PI3K inhibitor (wortmannin) on above-mentioned process was assayed. The results showed that thrombin elevated the activity of RhoA and the binding capability of RhoA with mDia1 during thrombin-induced platelet aggregation and spreading on Fg coated coverslips. Wortmannin inhibited the rising of RhoA activity and the binding level of RhoA with mDia1 induced by thrombin. Thrombin elevated the activity of Rac1 and Cdc42 during thrombin-induced platelet aggregation, but could not induce binding of Rac1 or Cdc42 with mDia1. Wortmannin could not inhibit the rising of Rac1 and Cdc42 activity induced by thrombin. It is concluded that the PI3-kinase regulates the thrombin-induced actin cytoskeleton reconstitution in platelets by RhoA-mDia1 pathway.
Actins ; metabolism ; pharmacology ; Adaptor Proteins, Signal Transducing ; immunology ; metabolism ; Blood Platelets ; metabolism ; Cells, Cultured ; Humans ; Phosphatidylinositol 3-Kinases ; pharmacology ; Platelet Aggregation ; drug effects ; Thrombin ; pharmacology ; rac1 GTP-Binding Protein ; metabolism ; rhoA GTP-Binding Protein ; metabolism ; pharmacology

The regulation mechanism of interferon (IFN) and IFN-stimulated genes is a very complex procedure and is dependent on cell types and virus species. We observed molecular changes related to anti-viral responses in endothelial cells during Hantaan virus (HTNV) infection. We found that there are two patterns of gene expression, the first pattern of gene expression being characterized by early induction and short action, as in that of type I IFNs,' and the other being characterized by delayed induction and long duration, as those of IRF-7, MxA, and TAP-1/2. Even though there are significant differences in their induction folds, we found that all of IFN-alpha/beta , IRF- 3/7, MxA, and TAP-1/2 mRNA expressions reached the peak when the viral replication was most active, which took place 3 days of post infection (d.p.i.). In addition, an interesting phenomenon was observed; only one gene was highly expressed in paired genes such as IFN-alpha/beta??(3/277-folds), IRF-3/7 (2.2/29.4-folds), and TAP- 1/2 (26.2/6.1-folds). Therefore, IFN-beta, IRF-7, and TAP-1 seem to be more important for the anti-viral response in HTNV infection. MxA was increased to 296-folds at 3 d.p.i. and kept continuing 207-folds until 7 d.p.i.. The above results indicate that IFN-beta works for an early anti-viral response, while IRF7, MxA, and TAP-1 work for prolonged anti-viral response in HTNV infection.
ATP-Binding Cassette Transporters/*genetics ; Blotting, Western ; Cells, Cultured ; Endothelial Cells/metabolism/*virology ; GTP-Binding Proteins/*genetics ; *Gene Expression Regulation ; Hantaan virus/*immunology ; Histocompatibility Antigens Class I/analysis ; Humans ; Interferon Regulatory Factor-3/genetics ; Interferon Regulatory Factor-7/*genetics ; Interferons/*genetics ; RNA, Messenger/analysis

OBJECTIVETo investigate the influence of activated complement on the secretory function of peritoneal macrophage (PMphi) in the production of nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha), especially in the role of different G-protein subtypes in this process after burns.

METHODSThe mice inflicted by 18% TBSA full-thickness scald was established and employed as the model. And the mice were divided into A (the complements were preserved and activated) and B (with intraperitoneal injection of CVF to deplete complement before scald) groups. The plasma of the mice in the two groups was collected at 6 postburn hour (PBH) and cultured with PMphi from normal mice. The PMphi were pretreated with pertussis toxin (PT) and with cholera toxin (CT). The NO and TNF-alpha levels in the supernatant of normal PMphi culture with different pretreatment were measured by Greiss assay.

RESULTSThe NO and TNF-alpha contents in group A [(80 +/- 12) micromol/L, (46 +/- 6)%] were obviously higher than those in group B [(34 +/- 5) micromol/L, (26 +/- 5)%, P < 0.01]. The NO content produced by PMphi (45 +/- 10 micromol/L) in A group decreased (P < 0.01), while the TNF-alpha activity (58 +/- 10)% increased by PT pretreatment (P < 0.05). On the contrary, the NO content produced by PMphi (105 +/- 18 micromol/L) in group A increased (P < 0.01), while the TNF-alpha activity (27 +/- 6)% decreased by CT pretreatment (P < 0.01).

CONCLUSIONThese results indicates that the secretory function of normal PMphi can be enhanced by complement activation after thermal injury, which might partly be due to the effect of activated complement components through complement receptor coupled G-protein. In the secretory function of complement stimulated Mphi, Gi protein has a major role in the production of NO, Gs protein is mainly involved in the secretion of TNF-alpha.

Animals ; Burns ; immunology ; metabolism ; Complement Activation ; Complement System Proteins ; metabolism ; Female ; GTP-Binding Proteins ; metabolism ; Macrophage Activation ; immunology ; Macrophages, Peritoneal ; secretion ; Male ; Mice ; Mice, Inbred Strains ; Nitric Oxide ; biosynthesis ; Signal Transduction ; Tumor Necrosis Factor-alpha ; biosynthesis