No abstract

Flaviviruses accumulate abundant subgenomic RNA (sfRNA) in infected cells. It has been reported that sfRNA results from stalling of host 5’-to-3’ exoribonuclease XRN1 at the highly structured RNA of the 3’ untranslated region (UTR). Although XRN1 digestion of a 3’-terminal 800-nt RNA could stall at a position to generate the sfRNA in vitro, we found that knocking out XRN1 had no effect on the accumulation of sfRNA in Japanese encephalitis virus (JEV) infected cells. Mutagenesis studies revealed that the stemloop II (SLII) at the 3’ UTR is required for the accumulation of sfRNA. According to the results of an in vitro RNA-dependent RNA polymerase (RdRp) assay, the (-)10431-10566 RNA fragment, containing the putative promoter on the antigenome for the sfRNA transcription, binds to RdRp protein and exhibits a strong promoter activity. Taken together, our results indicate that the JEV sfRNA could be transcribed initially and then be trimmed by XRN1 or other unidentified exoribonucleases.

MicroRNAs (miRNAs) play versatile roles in multiple biological processes. However, little is known about miRNA’s involvement in flavivirus persistent infection. Here, we used an miRNA array analysis of Japanese encephalitis virus (JEV)-infected cells to search for persistent infection-associated miRNAs in comparison to acute infection. Among all differentially expressed miRNAs, the miR-125b-5p is the most significantly increased one. The high level of miR-125b-5p in persistently JEV-infected cells was confirmed by Northern analysis and real-time quantitative polymerase chain reaction. As soon as the cells established a persistent infection, a significantly high expression of miR-125b-5p was readily observed. Transfecting excess quantities of a miR-125b-5p mimic into acutely infected cells reduced genome replication and virus titers. Host targets of miR125b-5p were analyzed by target prediction algorithms, and six candidates were confirmed by a dual-luciferase reporter assay. These genes were upregulated in the acutely infected cells and sharply declined in the persistently infected cells. The transfection of the miR125b-5p mimic reduced the expression levels of Stat3, Map2k7, and Triap1. Our studies indicated that miR-125b-5p targets both viral and host sequences, suggesting its role in coordinating viral replication and host antiviral responses. This is the first report to characterize the potential roles of miR-125b-5p in persistent JEV infections.

No abstract

The development of low-cost, ultrasound-guided focused ultrasound (USgFUS) treatment platforms is expected to advance the adoption of microbubble (MB)-mediated brain therapy by improving access to the technology. Our group has designed and fabricated clinical-scale transmit/receive phased array systems for MB-mediated USgFUS brain therapy. Acoustic field simulations were carried out to optimize array element placement, and transducer scaffolds were constructed using 3D printing techniques. These devices have been employed for skull computed tomography-array registration, 3D spatial mapping of MB activity in vivo through ex-vivo human skullcaps via noninvasive aberration correction methods, and we have harnessed this spatiotemporal cavitation information to calibrate exposure levels for safe volumetric blood-brain barrier opening. At higher exposure levels, we have demonstrated the ability of 3D MB imaging data to predict the tissue damage volume distributions induced during nonthermal brain ablation. Ultrafast processing of acoustic emissions data has been shown to uncover MB dynamics hidden by conventional whole-burst temporal averaging, and can inform temporal undersampling strategies when millisecond-long tone bursts are applied. Machine learning approaches can assist with image-based classification of MB activity, which may result in finer control of the induced bioeffects. This talk will focus on our recent results obtained with a novel modular USgFUS phased array system.