Development of an assay system for the analysis of host RISC activity in the presence of a potyvirus RNA silencing suppressor, HC-Pro.

BACKGROUND: To investigate the mechanism of RNA silencing suppression, the genetic transformation of viral suppressors of RNA silencing (VSRs) in Arabidopsis integrates ectopic VSR expression at steady state, which overcomes the VSR variations caused by different virus infections or limitations of host range. Moreover, identifying the insertion of the transgenic VSR gene is necessary to establish a model transgenic plant for the functional study of VSR. METHODS: Developing an endogenous AGO1-based in vitro RNA-inducing silencing complex (RISC) assay prompts further investigation into VSR-mediated suppression. Three P1/HC-Pro plants from turnip mosaic virus (TuMV) (P1/HC-ProTu), zucchini yellow mosaic virus (ZYMV) (P1/HC-ProZy), and tobacco etch virus (TEV) (P1/HC-ProTe) were identified by T-DNA Finder and used as materials for investigations of the RISC cleavage efficiency. RESULTS: Our results indicated that the P1/HC-ProTu plant has slightly lower RISC activity than P1/HC-ProZy plants. In addition, the phenomena are consistent with those observed in TuMV-infected Arabidopsis plants, which implies that HC-ProTu could directly interfere with RISC activity. CONCLUSIONS: In this study, we demonstrated the application of various plant materials in an in vitro RISC assay of VSR-mediated RNA silencing suppression.

Acemannan-induced tooth socket healing: A 12-month randomized controlled trial.

BACKGROUND/PURPOSE: Natural compounds have become alternatives for bone regeneration. Acemannan, the main polysaccharide extracted from Aloe vera, has been demonstrated as a promising osteoinductive material in vitro and in vivo. This clinical study investigated the effect of acemannan on tooth socket healing. MATERIALS AND METHODS: Thirty-five otherwise healthy patients, 18-25 years old and diagnosed with horizontal or vertical partial impaction of the lower third molars, were enrolled in this randomized controlled trial. After removing the teeth, the sockets randomly received one of the following treatments: spontaneous blood-clotting (control), 20 mg acemannan sponge, or 50 mg acemannan sponge. Cone-beam computed tomography of the mandible was performed immediately (baseline), and at 3-, 6-, and 12-months postoperatively; the data were analyzed using the OsiriX MD program. Bone healing in the socket was determined measuring the socket volume. One-way ANOVA was used to analyze the differences within each group and between groups. RESULTS: Thirty-five patients with 43 partially impacted lower third molars participated in this study. No patients exhibited alveolar osteitis or secondary infection. Compared with baseline, all groups showed significant reduction in socket volume at all observation time-points (p < 0.05). The 50 mg acemannan group had a significantly greater reduction in socket volume compared with the control at all postoperative time-points (p < 0.05). The 20 mg group had a significantly greater reduction in socket volume compared with the control at 3-months postoperatively (p < 0.05). CONCLUSION: We conclude that acemannan increases bone healing at 3-, 6-, and 12-months after removal of partially impacted mandibular third molars.

Investigation of P1/HC-Pro-Mediated ABA/Calcium Signaling Responses via Gene Silencing through High- and Low-Throughput RNA-seq Approaches.

The P1/HC-Pro viral suppressor of potyvirus suppresses posttranscriptional gene silencing (PTGS). The fusion protein of P1/HC-Pro can be cleaved into P1 and HC-Pro through the P1 self-cleavage activity, and P1 is necessary and sufficient to enhance PTGS suppression of HC-Pro. To address the modulation of gene regulatory relationships induced by turnip mosaic virus (TuMV) P1/HC-Pro (P1/HC-ProTu), a comparative transcriptome analysis of three types of transgenic plants (P1Tu, HC-ProTu, and P1/HC-ProTu) were conducted using both high-throughput (HTP) and low-throughput (LTP) RNA-Seq strategies. The results showed that P1/HC-ProTu disturbed the endogenous abscisic acid (ABA) accumulation and genes in the signaling pathway. Additionally, the integrated responses of stress-related genes, in particular to drought stress, cold stress, senescence, and stomatal dynamics, altered the expressions by the ABA/calcium signaling. Crosstalk among the ABA, jasmonic acid, and salicylic acid pathways might simultaneously modulate the stress responses triggered by P1/HC-ProTu. Furthermore, the LTP network analysis revealed crucial genes in common with those identified by the HTP network in this study, demonstrating the effectiveness of the miniaturization of the HTP profile. Overall, our findings indicate that P1/HC-ProTu-mediated suppression in RNA silencing altered the ABA/calcium signaling and a wide range of stress responses.