Black carp A20 inhibits interferon signaling through de-ubiquitinating IKKß.

IkappaB kinase beta (IKKß) is a key member of IκB kinases and functions importantly in interferon (IFN) signaling. Phosphorylation and ubiquitination are involved in the activation of IKKß. A20 is a de-ubiquitin enzyme and functions as a suppressor in inflammation signaling, which has been reported to be phosphorylated and activated by IKKß. However, the role and relationship of IKKß and A20 in teleost remains unclear. In this study, IKKß (bcIKKß) and A20 (bcA20) of black carp (Mylopharyngodon piceus) have been cloned and characterized. Overexpressed bcIKKß in EPC cells showed strong anti-viral ability by activating both NF-κB and IFN signaling. EPC cells stable expressing bcIKKß presented improved anti-viral activity as well. The interaction between bcA20 and bcIKKß was identified, and overexpression of bcA20 was able to suppress bcIKKß-mediated activation of NF-κB and IFN signaling. Meanwhile, knock-down of A20 increased host the antiviral ability of host cells. Importantly, it has been identified that bcA20 was able to remove K27-linked ubiquitination and decrease the phosphorylation of bcIKKß. Thus, our data conclude that bcA20 suppresses the anti-viral activity of bcIKKß and removes its K27-linked ubiquitination, which presents a new mechanism of IKKß regulation.

Knockdown of Stanniocalcin-1 inhibits growth and glycolysis in oral squamous cell carcinoma cells.

Oral squamous cell carcinoma (OSCC) is the most common malignancy among head and neck squamous cell carcinomas. Targeted therapy plays a crucial role in the treatment of OSCC. However, new and more targets are still needed to develop. Stanniocalcin-1 (STC-1) is a glycoprotein hormone that affects the progression of cancers. However, the potential role of STC-1 in OSCC progression remains to be explored. Here, we aimed to elucidate the role of STC-1 in OSCC. We revealed that STC-1 was highly expressed in OSCC tissues and is correlated with poor patient prognosis. Knockdown of STC-1 significantly suppressed the growth of OSCC cells and restrained glycolysis by reducing glucose consumption, ATP production, and lactate levels. Mechanistically, STC-1 ablation inhibited the PI3K/Akt pathway, reducing the phosphorylation levels of PI3K and Akt. In conclusion, STC-1 depletion restrained OSCC cell growth and glycolysis via PI3K/Akt pathway and has the potential to serve as a therapeutic target for OSCC.

IFNb of black carp functions importantly in host innate immune response as an antiviral cytokine.

Type I interferons (IFN-Is) play an important role in the antiviral immune response in teleost fishes. In this study, one type I interferon (bcIFNb) from black carp (Mylopharyngodon piceus) has been cloned and characterized. The full-length cDNA of bcIFNb gene consists of 806 nucleotides and the predicted bcIFNb protein contains 188 amino acids. Basing on the cysteine number and evolutionary position, bcIFNb was classified into group II type I IFN. q-PCR analysis demonstrated that bcIFNb mRNA level varied in vivo and ex vivo in response to different stimuli. bcIFNb was detected in both the whole cell lysate and the supernatant media of HEK293T cells or EPC cells transfected with bcIFNb through immunoblot assay. IFN stimulated genes (ISGs) were greatly upregulated when the host cells were treated with the bcIFNb-containing conditioned media. EPC cells showed greatly enhanced antiviral ability when the cells were transfected with bcIFNb or treated with the bcIFNb-containing conditioned media before GCRV or SVCV infection. Glycosidase digestion analysis determined that bcIFNb was modified with N-linked glycosylation, which occurred on the Asn (N) of 92 site of this cytokine. The un-glycosylated mutant bcIFNb-N92Q presented the similar antiviral ability as that of wild type bcIFNb, which demonstrated that N-linked glycosylation did not contribute directly to the antiviral property of this fish cytokine.

TRAF2 of black carp upregulates MAVS-mediated antiviral signaling during innate immune response.

Tumor necrosis factor receptor-associated factor 2 (TRAF2) is a crucial component of the tumor necrosis factor (TNF) mediated signaling of higher vertebrates. To elucidate its function in teleost fish, TRAF2 homologue of black carp (Mylopharyngodon piceus) has been cloned and characterized in this study. The open reading frame (ORF) of black carp TRAF2 (bcTRAF2) consists of 1611 nucleotides and bcTRAF2 contains 536 amino acids. bcTRAF2 protein migrated around 65 KDa in immunoblot analysis of both EPC and HEK293T cells. bcTRAF2 was identified as a cytosolic protein and suggested to be associated with vesicles scattering in the cytoplasm. NF-κB transcription instead of IFN transcription was activated by bcTRAF2 in reporter assay. It was interesting that bcMAVS-mediated IFN production was up-regulated by bcTRAF2 in a dose dependent manner in reporter assay. Accordingly, EPC cells transfected with both bcMAVS and bcTRAF2 showed enhanced antiviral activity comparing EPC cells only expressing bcMAVS. When co-expressed with bcMAVS, bcTRAF2 was redistributed in the cytoplasm and its subcellular location overlapped with the subcellular location of bcMAVS, which suggested the association between these two molecules. Taken together, the data generated in this paper supported the conclusion that bcTRAF2 was recruited into host innate immune response and positively regulated bcMAVS-mediated antiviral signaling.

Characterization of the black carp TRAF6 signaling molecule in innate immune defense.

Tumor necrosis factor receptor-associated factor 6 (TRAF6) plays a vital role in the innate immune response of higher vertebrates. To elucidate its function in teleost fish, TRAF6 homologue of black carp (Mylopharyngodon piceus) has been cloned and characterized in this study. Black carp TRAF6 (bcTRAF6) transcription in Mylopharyngodon piceus fin (MPF) cells was up-regulated in response to both poly (I:C) treatment and viral infection, but was suppressed by LPS stimulation. bcTRAF6 migrated around 72 KDa in immunoblot analysis and was identified as a cytosolic protein suggested to be associated with vesicles scattering in the cytoplasm. Reporter assay demonstrated that NF-κB instead of IFN was activated by bcTRAF6; and EPC cells expressing bcTRAF6 presented the same cytopathic effect (CPE) ratio to that of control cells. When co-expressed with bcMAVS, bcTRAF6 was redistributed and overlapped with the subcellular location of bcMAVS. It was interesting that bcMAVS mediated the IFN induction was up-regulated by low input of bcTRAF6 but down-regulated by high input of bcTRAF6. Taken together, the data generated in this paper supported the conclusion that bcTRAF6 associated with bcMAVS and was recruited into bcMAVS mediated signaling during host innate immune response.

Construction of three-dimensional graphene interfaces into carbon fiber textiles for increasing deposition of nickel nanoparticles: flexible hierarchical magnetic textile composites for strong electromagnetic shielding.

Since manipulating electromagnetic waves with electromagnetic active materials for environmental and electric engineering is a significant task, here a novel prototype is reported by introducing reduced graphene oxide (RGO) interfaces in carbon fiber (CF) networks for a hierarchical carbon fiber/reduced graphene oxide/nickel (CF-RGO-Ni) composite textile. Upon charaterizations of the microscopic morphologies, electrical and magnetic properties, the presence of three-dimensional RGO interfaces and bifunctional nickel nanoparticles substantially influences the related physical properties in the resulting hierarchical composite textiles. Eletromagnetic interference (EMI) shielding performance suggests that the hierarchical composite textiles hold a strong shielding effectiveness greater than 61 dB, showing greater advantages than conventional polymeric and foamy shielding composites. As a polymer-free lightweight structure, flexible CF-RGO-Ni composites of all electromagnetic active components offer unique understanding of the multi-scale and multiple mechanisms in electromagnetic energy consumption. Such a novel prototype of shielding structures along with convenient technology highlight a strategy to achieve high-performance EMI shielding, coupled with a universal approach for preparing advanced lightweight composites with graphene interfaces.