Image_3_Stimulation of the Internal Ribosome Entry Site (IRES)-Dependent Translation of Enterovirus 71 by DDX3X RNA Helicase and Viral 2A and 3C Proteases.JPEG

<p>The translation of enterovirus 71 (EV71) is mediated by an internal ribosome entry site (IRES)-dependent manner. EV71 IRES comprises five highly structured domains (domains II-VI) in the 5′-untranslated region of the viral mRNA. A conserved AUG triplet residing in domain VI is proposed to be the ribosome entry site. It is thus envisaged that the highly structured conformation of domain VI may actually reduce the accessibility of the AUG triplet to the ribosome. This study identified a DEAD-box family RNA helicase, DDX3X, that positively regulated the EV71 IRES-dependent translation. The helicase activity of DDX3X was required for the stimulation of EV71 IRES activity; however, DDX3X was no longer important for the IRES activity when the secondary structure of domain VI was destabilized. DDX3X interacted with the truncated eIF4G which bound specifically to domain V. Thus, we proposed that DDX3X might bind to domain VI or a region nearby via the interaction with the truncated eIF4G, and subsequently unwound the secondary structure of domain VI to facilitate ribosome entry. Additionally, we demonstrated that the viral 2A^pro and 3C^pro enhanced the IRES-dependent translation via their protease activities. Together, these results indicate that DDX3X is an important RNA helicase involved in EV71 IRES-dependent translation and that IRES translation is enhanced by viral infection, partly mediated by viral protease activity.</p

Image_1_Stimulation of the Internal Ribosome Entry Site (IRES)-Dependent Translation of Enterovirus 71 by DDX3X RNA Helicase and Viral 2A and 3C Proteases.JPEG

<p>The translation of enterovirus 71 (EV71) is mediated by an internal ribosome entry site (IRES)-dependent manner. EV71 IRES comprises five highly structured domains (domains II-VI) in the 5′-untranslated region of the viral mRNA. A conserved AUG triplet residing in domain VI is proposed to be the ribosome entry site. It is thus envisaged that the highly structured conformation of domain VI may actually reduce the accessibility of the AUG triplet to the ribosome. This study identified a DEAD-box family RNA helicase, DDX3X, that positively regulated the EV71 IRES-dependent translation. The helicase activity of DDX3X was required for the stimulation of EV71 IRES activity; however, DDX3X was no longer important for the IRES activity when the secondary structure of domain VI was destabilized. DDX3X interacted with the truncated eIF4G which bound specifically to domain V. Thus, we proposed that DDX3X might bind to domain VI or a region nearby via the interaction with the truncated eIF4G, and subsequently unwound the secondary structure of domain VI to facilitate ribosome entry. Additionally, we demonstrated that the viral 2A^pro and 3C^pro enhanced the IRES-dependent translation via their protease activities. Together, these results indicate that DDX3X is an important RNA helicase involved in EV71 IRES-dependent translation and that IRES translation is enhanced by viral infection, partly mediated by viral protease activity.</p

Image_2_Stimulation of the Internal Ribosome Entry Site (IRES)-Dependent Translation of Enterovirus 71 by DDX3X RNA Helicase and Viral 2A and 3C Proteases.JPEG

<p>The translation of enterovirus 71 (EV71) is mediated by an internal ribosome entry site (IRES)-dependent manner. EV71 IRES comprises five highly structured domains (domains II-VI) in the 5′-untranslated region of the viral mRNA. A conserved AUG triplet residing in domain VI is proposed to be the ribosome entry site. It is thus envisaged that the highly structured conformation of domain VI may actually reduce the accessibility of the AUG triplet to the ribosome. This study identified a DEAD-box family RNA helicase, DDX3X, that positively regulated the EV71 IRES-dependent translation. The helicase activity of DDX3X was required for the stimulation of EV71 IRES activity; however, DDX3X was no longer important for the IRES activity when the secondary structure of domain VI was destabilized. DDX3X interacted with the truncated eIF4G which bound specifically to domain V. Thus, we proposed that DDX3X might bind to domain VI or a region nearby via the interaction with the truncated eIF4G, and subsequently unwound the secondary structure of domain VI to facilitate ribosome entry. Additionally, we demonstrated that the viral 2A^pro and 3C^pro enhanced the IRES-dependent translation via their protease activities. Together, these results indicate that DDX3X is an important RNA helicase involved in EV71 IRES-dependent translation and that IRES translation is enhanced by viral infection, partly mediated by viral protease activity.</p

sj-docx-1-nnr-10.1177_15459683231170539 – Supplemental material for Effects of Neurofeedback on Cognitive Function, Productive Activity, and Quality of Life in Patients With Traumatic Brain Injury: A Randomized Controlled Trial

Supplemental material, sj-docx-1-nnr-10.1177_15459683231170539 for Effects of Neurofeedback on Cognitive Function, Productive Activity, and Quality of Life in Patients With Traumatic Brain Injury: A Randomized Controlled Trial by Pin-Yuan Chen, I-Chang Su, Chun-Ying Shih, Yen-Chun Liu, Yu-Kai Su, Li Wei, Hui-Tzung Luh, Hui-Chuan Huang, Pei-Shan Tsai, Yen-Chun Fan and Hsiao-Yean Chiu in Neurorehabilitation and Neural Repair</p

Construction of a Near-Infrared-Activatable Enzyme Platform To Remotely Trigger Intracellular Signal Transduction Using an Upconversion Nanoparticle

Photoactivatable (caged) bioeffectors provide a way to remotely trigger or disable biochemical pathways in living organisms at a desired time and location with a pulse of light (uncaging), but the phototoxicity of ultraviolet (UV) often limits its application. In this study, we have demonstrated the near-infrared (NIR) photoactivatable enzyme platform using protein kinase A (PKA), an important enzyme in cell biology. We successfully photoactivated PKA using NIR to phosphorylate its substrate, and this induced a downstream cellular response in living cells with high spatiotemporal resolution. In addition, this system allows NIR to selectively activate the caged enzyme immobilized on the nanoparticle surface without activating other caged proteins in the cytosol. This NIR-responsive enzyme–nanoparticle system provides an innovative approach to remote-control proteins and enzymes, which can be used by researchers who need to avoid direct UV irradiation or use UV as a secondary channel to turn on a bioeffector

Up-Regulation of MicroRNA-190b Plays a Role for Decreased IGF-1 That Induces Insulin Resistance in Human Hepatocellular Carcinoma

<div><p>Background & Aims</p><p>Insulin-like growth factor, (IGF)-1, is produced mainly by the liver and plays important roles in promoting growth and regulating metabolism. Previous study reported that development of hepatocellular carcinoma (HCC) was accompanied by a significant reduction in serum IGF-1 levels. Here, we hypothesized that dysregulation of microRNAs (miRNA) in HCC can modulate IGF-1 expression post-transcriptionally.</p><p>Methods</p><p>The miRNAs expression profiles in a dataset of 29 HCC patients were examined using illumina BeadArray. Specific miRNA (miR)-190b, which was significantly up-regulated in HCC tumor tissues when compared with paired non-tumor tissues, was among those predicted to interact with 3′-untranslated region (UTR) of <i>IGF-1.</i> In order to explore the regulatory effects of miR-190b on IGF-1 expression, luciferase reporter assay, quantitative real-time PCR, western blotting and immunofluorecence analysis were performed in HCC cells.</p><p>Results</p><p>Overexpression of miR-190b in Huh7 cells attenuated the expression of IGF-1, whereas inhibition of miR-190b resulted in up-regulation of IGF-1. Restoration of IGF-1 expression reversed miR-190b-mediated impaired insulin signaling in Huh7 cells, supporting that IGF-1 was a direct and functional target of miR-190b. Additionally, low serum IGF-1 level was associated with insulin resistance and poor overall survival in HCC patients.</p><p>Conclusions</p><p>Increased expression of miR-190 may cause decreased IGF-1 in HCC development. Insulin resistance appears to be a part of the physiopathologic significance of decreased IGF-1 levels in HCC progression. This study provides a novel miRNA-mediated regulatory mechanism for controlling IGF-1 expression in HCC and elucidates the biological relevance of this interaction in HCC.</p></div

The regulatory effects of miR-190b on IGF-1 expression.

<p>(A) qRT-PCR assay verified the relative miR-190b expression in Huh7 stable cells clones. (B&C) Overexpression of miR-190b attenuated IGF-1 protein expression, and blocking miR-190b moderately increased IGF-1 protein expression. These results were assayed by western blotting (B) and immunofluorescence staining (C), respectively. (D) IGF-1 mRNA was detected by qRT-PCR in stable cells expressing precursor miR-190b or inhibitor miR-190b. Relative expression of IGF-1 compared to negative control (NC) was calculated using the 2^−ΔΔCT methods. (E) The endogenous transcript levels of miR-190b and IGF-1 were dertermined by qRT-PCR in Huh7 and HepG2 cells. (F&G) qRT-PCR analysis of miR-190b (F) and IGF-1 (G) in HepG2 cells transfected with anti-NC or anti-miR-190b vectors. (H) Western blotting of IGF-1 in HepG2 cells transfected with anti-NC or anti-miR-190b vectors. Relative intensity (RI) shown was calculated by normalization of the intensities of IGF-1 from the internal controls. **, <i>P</i><0.01 and *, <i>P</i><0.05 as compared to each of the negative control cells.</p

Over-expression of miR-190b in HCC cells leads to insulin resistance through down-regulation of IGF-1 expression.

<p>(A) Representative western blotting and quantification of expression and insulin-stimulated phosphorylation of IRS1, FOXO1 and GSK3β in Huh7 stable cells expressing precursor negative control (NC) or miR-190b. Cells were starved for 24 hours and then treated with 100 nM insulin for 10 and 30 minutes. Band intensity and ratio of phospho-form/total proteins were calculated by Image J. (B) immunofluorescence staining showing the subcellular localization of FOXO1 in Huh7 stable cells. (C) Extracellular glucose production in Huh7 stable cells was measured as described in Materials and Methods (*<i>P</i><0.05).</p

IGF-1 was a direct target of miR-190b.

<p>(A) Base pairing complement suggests two putative miR-190b binding positions at 844–850 and 1752–1758 of the IGF-1 3′UTR. (B) The effect of precursor miR-190b or miR-190b inhibitor on pEZX-IGF-1 3′UTR luciferase activity. Luciferase assays were done on HEK293 T cells. Significant differences were compared to each negative control (NC). (C) HEK293T cells were transiently transfected with pre-miR-190b or pre-NC vector together with the pEZX-MT01 empty vector, a modified pEZX-MT01 vector containing wild-type IGF-1 3′UTR, or two mutant IGF-1 3′UTR carrying mutations for each putative miR-190b binding site. Luciferase activity of pEZX-MT01 empty plasmid was set to 1. ***, <i>P</i><0.001; **, <i>P</i><0.01; *, <i>P</i><0.05.</p

Low IGF-1 is associated with insulin resistance and poor prognosis in HCC patients.

<p>(A, B) Kaplan-Meier curves for overall survival (A) and recurrence-free survival (B) in patients with HCC after curative resection. Compared with patients with high IGF-1 levels, patients with low IGF-1 levels exhibited significantly shorter overall survival but no significant difference in recurrence-free survival. Statistical significance was calculated using the log-rank test. (C, D) Correlations of insulin levels (C) and homeostasis model assessment of insulin resistance (HOMA-IR) (D) with serum IGF-1 levels in HCC patients. Linear regression coefficients and statistical significance are indicated.</p