TALANTA
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2019
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IF 4.916
Zengjin Li,Hui Guo,Fangyuan Xu,Wei Tang,Xinrui Duan
We reported a new strategy for sensitive monitoring in vitro RNA synthesis in real time based on fluorescence resonance energy transfer (FRET) from water-soluble conjugated polymer poly (9, 9-bis (6′-N, N, N,-trimethylammonium) hexyl) fluorene-co-alt-1,4-phenylene) bromide (PFP) to fluorogenic RNA aptamer/fluorophore (Spanich2/DFHBI and Broccoli/DFHBI) system. In this strategy, RNA of interest was transcribed accompanied by the Spanich2 or Broccoli. Then the 3, 5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI) bound to the RNA aptamer sequence and thereby induced a fluorescence signal. PFP was used as the fluorescence energy donor, and Spanich2/DFHBI was the fluorescence energy acceptor. The fluorescence signal of Spanich2/DFHBI was amplified by light-harvesting and fluorescence amplification ability of PFP via FRET. And the limit of detection (LOD) (0.29 nM) was near 10-fold lower than that of RNA aptamer/DFHBI (LOD is 2.8 nM) alone by measuring the FRET ratio, which greatly reduced the variation of background signals. Most importantly, the addition of PFP did not interfere with RNA transcription in vitro , so this method was successfully applied to sensitively monitor RNA transcription and effect of T7 RNA polymerase inhibitor in real time, supplying a sensitive and simple method to study the modulation and inhibitor of RNA polymerase in vitro .
研究领域:
生物化学
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分子生物学
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分析化学
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DOI
PubMed
高影响因子
Scientific Reports
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2024
·Vol. 14
·期 1
·1-12
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IF 3.8
Hu Deliang,Yang Yuqing,Fang Lei,Fan Shijie,Lin Ling,Luo Puying,Xiong Yuanhuan,Su Yufang
Isoliquiritigenin (ISL), a naturally occurring flavonoid derived from licorice root, exhibits antioxidant, anticancer, anti-inflammatory, and anti-allergic properties, and is frequently detected in both environmental and human samples. Previous studies from our lab have demonstrated that ISL exposure can lead to developmental deformities and aberrant immune responses. However, the molecular mechanisms underlying ISL toxicity in zebrafish embryos remain incompletely elucidated. Therefore, this study aimed to elucidate the effects of ISL exposure on endoplasmic reticulum (ER) stress in zebrafish embryos by assessing the expression levels of ER stress markers HSPA5 and CHOP , along with associated apoptosis factors, under various ISL concentrations, with tunicamycin (TM) serving as a positive control. Furthermore, targeted analyses of ER stress-related pathways were conducted using RNA transcriptome sequencing, and the up-regulated gene was verified by western blot. The results revealed that ISL exposure significantly elevated the expression levels of HSPA5 and CHOP , concomitantly activating ER stress pathways, including pPERK-eIF2α-ATF4 and ATF6 pathways in zebrafish embryos. These findings suggest that the activation of endoplasmic reticulum stress signaling pathways may contribute to the developmental deformities observed in zebrafish embryos following ISL exposure, thereby highlighting the potential ecological risks associated with ISL usage.
研究领域:
毒理学
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发育生物学
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分子生物学
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环境科学
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PubMed
高影响因子
Journal of the American Chemical Society
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2025
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IF 15.6
Yiming Liu,Mingrui Zhang,Jingjing Zhao,Yongshuo Ren,Shubin Li,Weichen Wang,Wei Mu,Xiaojun Han
The reconstitution of the de novo nucleotide synthesis pathway from inorganic compounds for RNA transcription is essential for an autonomous artificial cell. Here, a de novo uridine triphosphate (UTP) synthesis pathway is successfully constructed starting from NH4HCO3, involving eight enzymes of carbamoyl phosphate synthetase (CPS), aspartate transcarbamoylase (ATC), dihydroorotase (DHO), dihydroorotate dehydrogenase (DHODH), ribose-phosphate pyrophosphokinase (RPPK), uridine 5′-monophosphate synthase (UMPs), uridylate kinase (UK), and nucleoside diphosphate kinase (NDK). ATP regeneration system composed of creatine kinase (CK) and creatine phosphate is incorporated to drive this pathway. 0.85 mM UTP is produced within 180 min under the optimized conditions, which is further used for RNA transcription along with preadded CTP, GTP, regenerated ATP, and T7 RNA polymerase. The de novo UTP synthesis pathway and RNA transcription components are encapsulated into giant unilamellar vesicles to build artificial cells capable of nucleotide de novo production for subsequent RNA transcription. The successful RNA transcription inside artificial cells is visualized using pyronin Y. The successful reconstitution of the de novo UTP synthesis pathway for RNA transcription inside artificial cells lays a solid foundation for the reproduction of an autonomous artificial cell.
研究领域:
合成生物学
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生物化学
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分子生物学
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人工细胞
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系统生物学
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DOI
PubMed
高影响因子
Bioactive Materials
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2024
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IF 18
Ying Jin,Hang Liu,Lei Chu,Jin Yang,Xiuyang Li,Hang Zhou,Haitao Jiang,Lei Shi,Jason Weeks,Joshua Rainbolt,Changjiang Yang,Thomas Xue,Haobo Pan,Zhongliang Deng,Chao Xie,Xu Cui,Youliang Ren
Implant-associated Staphylococcus aureus ( S. aureus ) osteomyelitis is a severe challenge in orthopedics. While antibiotic-loaded bone cement is a standardized therapeutic approach for S. aureus osteomyelitis, it falls short in eradicating Staphylococcus abscess communities (SACs) and bacteria within osteocyte-lacuna canalicular network (OLCN) and repairing bone defects. To address limitations, we developed a borosilicate bioactive glass (BSG) combined with ferroferric oxide (Fe 3 O 4 ) magnetic scaffold to enhance antibacterial efficacy and bone repair capabilities. We conducted comprehensive assessments of the osteoinductive, immunomodulatory, antibacterial properties, and thermal response of this scaffold, with or without an alternating magnetic field (AMF). Utilizing a well-established implant-related S. aureus tibial infection rabbit model, we evaluated its antibacterial performance in vivo . RNA transcriptome sequencing demonstrated that BSG + 5%Fe 3 O 4 enhanced the immune response to bacteria and promoted osteogenic differentiation and mineralization of MSCs. Notably, BSG + 5%Fe 3 O 4 upregulated gene expression of NOD-like receptor and TNF pathway in MSCs, alongside increased the expression of osteogenic factors (RUNX2, ALP and OCN) in vitro . Flow cytometry on macrophage exhibited a polarization effect towards M2, accompanied by upregulation of anti-inflammatory genes (TGF-β1 and IL-1Ra) and downregulation of pro-inflammatory genes (IL-6 and IL-1β) among macrophages. In vivo CT imaging revealed the absence of osteolysis and periosteal response in rabbits treated with BSG + 5%Fe 3 O 4 + AMF at 42 days. Histological analysis indicated complete controls of SACs and bacteria within OLCN by day 42, along with new bone formation, signifying effective control of S. aureus osteomyelitis. Further investigations will focus on the in vivo biosafety and biological mechanism of this scaffold within infectious microenvironment.
研究领域:
骨科
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生物材料
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感染性疾病
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分子生物学
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生物医学工程
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DOI
PubMed
高影响因子
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