ZIKV infection activates the IRE1-XBP1 and ATF6 pathways of unfolded protein response in neural cells.

BACKGROUND: Many viruses depend on the extensive membranous network of the endoplasmic reticulum (ER) for their translation, replication, and packaging. Certain membrane modifications of the ER can be a trigger for ER stress, as well as the accumulation of viral protein in the ER by viral infection. Then, unfolded protein response (UPR) is activated to alleviate the stress. Zika virus (ZIKV) is a mosquito-borne flavivirus and its infection causes microcephaly in newborns and serious neurological complications in adults. Here, we investigated ER stress and the regulating model of UPR in ZIKV-infected neural cells in vitro and in vivo. METHODS: Mice deficient in type I and II IFN receptors were infected with ZIKV via intraperitoneal injection and the nervous tissues of the mice were assayed at 5 days post-infection. The expression of phospho-IRE1, XBP1, and ATF6 which were the key markers of ER stress were analyzed by immunohistochemistry assay in vivo. Additionally, the nuclear localization of XBP1s and ATF6n were analyzed by immunohistofluorescence. Furthermore, two representative neural cells, neuroblastoma cell line (SK-N-SH) and astrocytoma cell line (CCF-STTG1), were selected to verify the ER stress in vitro. The expression of BIP, phospho-elF2α, phospho-IRE1, and ATF6 were analyzed through western blot and the nuclear localization of XBP1s was performed by confocal immunofluorescence microscopy. RT-qPCR was also used to quantify the mRNA level of the UPR downstream genes in vitro and in vivo. RESULTS: ZIKV infection significantly upregulated the expression of ER stress markers in vitro and in vivo. Phospho-IRE1 and XBP1 expression significantly increased in the cerebellum and mesocephalon, while ATF6 expression significantly increased in the mesocephalon. ATF6n and XBP1s were translocated into the cell nucleus. The levels of BIP, ATF6, phospho-elf2α, and spliced xbp1 also significantly increased in vitro. Furthermore, the downstream genes of UPR were detected to investigate the regulating model of the UPR during ZIKV infection in vitro and in vivo. The transcriptional levels of atf4, gadd34, chop, and edem-1 in vivo and that of gadd34 and chop in vitro significantly increased. CONCLUSION: Findings in this study demonstrated that ZIKV infection activates ER stress in neural cells. The results offer clues to further study the mechanism of neuropathogenesis caused by ZIKV infection

Zika virus promotes CCN1 expression via the CaMKIIα-CREB pathway in astrocytes.

Zika virus (ZIKV) infection in the human central nervous system (CNS) causes Guillain-Barre syndrome, cerebellum deformity, and other diseases. Astrocytes are immune response cells in the CNS and an important component of the blood-brain barrier. Consequently, any damage to astrocytes facilitates the spread of ZIKV in the CNS. Connective tissue growth factor/Nephroblastoma overexpressed gene family 1 (CCN1), an important inflammatory factor secreted by astrocytes, is reported to regulate innate immunity and viral infection. However, the mechanism by which astrocyte viral infection affects CCN1 expression remains undefined. In this study, we demonstrate that ZIKV infection up-regulates CCN1 expression in astrocytes, thus promoting intracellular viral replication. Other studies revealed that the cAMP response element (CRE) in the CCN1 promoter is activated by the ZIKV NS3 protein. The cAMP-responsive element-binding protein (CREB), a transacting factor of the CRE, is also activated by NS3 or ZIKV. Furthermore,a specific inhibitor of CREB, i.e. SGC-CBP30, reduced ZIKV-induced CCN1 up-regulation and ZIKV replication. Moreover, co-immunoprecipitation, overexpression, and knockdown studies confirmed that the interaction between NS3 and the regulatory domain of CaMKIIα could activate the CREB pathway, thus resulting in the up-regulation of CCN1 expression and enhancement of virus replication. In conclusion, the findings of our investigations on the NS3-CaMKIIα-CREB-CCN1 pathway provide a foundation for understanding the infection mechanism of ZIKV in the CNS

Discussion on Event Horizon and Quantum Ergosphere of Evaporating Black Holes in a Tunnelling Framework

In this paper, with the Parikh-Wilczek tunnelling framework the positions of the event horizon of the Vaidya black hole and the Vaidya-Bonner black hole are calculated respectively. We find that the event horizon and the apparent horizon of these two black holes correspond respectively to the two turning points of the Hawking radiation tunnelling barrier. That is, the quantum ergosphere coincides with the tunnelling barrier. Our calculation also implies that the Hawking radiation comes from the apparent horizon.Comment: 8 page

Tunnelling Effect and Hawking Radiation from a Vaidya Black Hole

In this paper, we extend Parikh' work to the non-stationary black hole. As an example of the non-stationary black hole, we study the tunnelling effect and Hawking radiation from a Vaidya black hole whose Bondi mass is identical to its mass parameter. We view Hawking radiation as a tunnelling process across the event horizon and calculate the tunnelling probability. We find that the result is different from Parikh's work because $\frac{dr_{H}}{dv}$ is the function of Bondi mass m(v)

The influence of aerobic sludge retention time on anaerobic co-digestion of dyeing and printing wastewater and sewage sludge

Two pilot-scale activated sludge systems comprising anaerobic baffled reactor (ABR) and aerobic plug flow reactor (PFR) were operated aiming to minimize excess sludge output of the activated sludgeprocess through coupled alkaline hydrolysis and anaerobic digestion. Variations in the effluent total chemical oxygen demand (TCOD) and NH4 +-N concentration proved that the process not only couldminimize excess sludge production but also guarantee the effluent TCOD well below the discharging limit (150 mg/l) if the inverse ratio of the aerobic sludge recirculation to anaerobic reactor did notexceed 60% for system A with 10 d aerobic sludge retention time (SRT) and 40% for system B (SRT was 25 d). The sludge activity at aerobic SRT of 25 d was evidently lower than aerobic SRT of 10 d. Those differences of sludge characteristics affected the inverse sludge ratio obviously. Aerobic bacteria after internal and external decay were converted to anoxic or anaerobic biomass. The distinct differences insludge yield of aerobic and anaerobic/anoxic processes could explain how aerobic SRT decided excess sludge activity which consequently affected anaerobic codigestion of printing and dyeing wastewaterand sewage sludge