N6-methyladenosine (m 6 A) is rapidly being studied and uncovered to play a significant role in various biological processes as well as in RNA fate and functions, while the effects of defined m 6 A sites are rarely characterized for the lack of convenient tools. To provide an applicable method to remove m 6 A modification at specific loci, an m 6 A editing system called "targeted RNA demethylation by SunTag system (TRADES)" is engineered. In this system, the targeting element dCas13b is fused to ten copies of GCN4 peptides which can recruit multiple scFv-fusion RNA demethylase to demethylate the target m 6 A site. Owing to this design, TRADES is more flexible to the indistinct m 6 A sites for its wide editing window. By site-specific demethylation of messenger RNA (mRNA) SON A2699, the lifetime of SON RNA is successfully prolonged in HeLa cells. Meanwhile, TRADES negligibly influences the lifetime of other non-targeted transcripts. TRADES also can regulate the gene expression of target transcript in an m 6 A-dependent manner. Moreover, the interference occuring for HBV and HIV replications demonstrates that the TRADES system holds potential in viral life cycle regulation and clinical applications.

Viruses have been shown to be equipped with regulatory RNAs to evade host defense system. It has long been known that rabies virus (RABV) transcribes a small regulatory RNA, leader RNA (leRNA), which mediates the transition from viral RNA transcription to replication. However, the detailed molecular mechanism remains enigmatic. In the present study, we determined the genetic architecture of RABV leRNA and demonstrated its inhibitory effect on replication of wild-type rabies, DRV-AH08. The RNA immunoprecipitation results suggest that leRNA inhibits RABV replication via interfering the binding of RABV nucleoprotein with genomic RNA. Furthermore, we identified heat shock cognate 70 kDa protein (Hsc70) as a leRNA host cellular interacting protein, of which the expression level was dynamically regulated by RABV infection. Notably, our data suggest that Hsc70 was involved in suppressing RABV replication by leader RNA. Finally, our experiments imply that leRNA might be potentially useful as a novel drug in rabies post-exposure prophylaxis. Together, this study suggested leRNA in concert with its host interacting protein Hsc70, dynamically down-regulate RABV replication.

Neurotropic viruses, such as the rabies virus (RABV) and Japanese encephalitis virus (JEV), induce neuronal dysfunction and complication, causing neuronal damage. Currently, there are still no effective clinical treatments for neuronal injury caused by neurotropic viruses. Memantine, a drug capable of passing through the blood‐brain barrier, noncompetitively and reversibly binds to n‐methyl‐ d‐aspartic acid (NMDA) receptors. Memantine is used to treat Alzheimer's disease by blocking the activation of extra axonal ion channels, thus preventing neuronal degeneration by inhibiting the abnormal cytosolic Ca 2+ increase. To explore whether memantine can alleviate neurological disturbances caused by RABV and JEV, the following experiments were carried out: (1) for primary neurons cultured in vitro infected with RABV, the addition of memantine showed neuroprotection. (2) In the RABV challenge experiments, memantine had limited therapeutic effect, mildly extending the survival time of mice. In contrast, memantine significantly prolonged the survival time of mice infected with JEV, by reducing the intravascular cuff and inflammatory cell infiltration in mice. Furthermore, memantine decreases the amount of JEV virus in mice brain.

Tuberculosis (TB) is the leading infectious cause of mortality worldwide, especially in developing countries. However, effective means for TB diagnosis, especially for bacillus-negative (Bn) TB laboratory diagnosis, are urgently needed. In the present study, serum IgG from each tuberculosis patients and healthy controls was purified using affinity chromatography. The samples were then analyzed using mass spectrometry (MS) and ultraperformance liquid chromatography (UPLC) methods. We quantitatively assessed the changes of serum IgG galactosylation in 567 human serum samples including 377 pulmonary TB patients and 190 healthy donors (HDs). We found significantly more agalactosylated (G0) vs monogalactosylated (G1) and digalactosylated (G2) N-glycans of IgG in TB patients, including smear-negative TB patients, than in HDs. The detection rate of TB diagnostic performance by MS for IgG–Gal ratio G0/(G1 + G2 × 2) is 90.48% for bacillus-positive (Bp) and 73.16% for Bn TB patients. Further, combination of MS method with other routine laboratory TB diagnostic methods significantly increased the detection rate to 91.01%–98.39%. Similar results were observed in Mycobacterium tuberculosis (M. tb) infection mouse models. The decrease in galactosylation of IgG in TB patients was also qualitatively confirmed using specific lectin blot assay. Using the above techniques, we can discriminate the content of IgG G0 with terminal N-acetylglucosamine and IgG–Gal ratio G0/(G1 + G2 × 2) between TB patients and HDs. Our data suggest that quantitative analysis of serum-based IgG–Gal ratio G0/(G1 + G2 × 2) could be used for TB auxiliary diagnosis with high effectiveness and feasibility and its combination with other routine laboratory TB diagnostic methods could remarkably improve the detection rate.

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