A Preliminary Survey of Transfer RNA Modifications and Modifying Enzymes of the Tropical Plant Cocos nucifera L.

The coconut (Cocos nucifera L.) is a commercial crop widely distributed among coastal tropical regions. It provides millions of farmers with food, fuel, cosmetics, folk medicine, and building materials. Among these, oil and palm sugar are representative extracts. However, this unique living species of Cocos has only been preliminarily studied at molecular levels. Benefiting from the genomic sequence data published in 2017 and 2021, we investigated the transfer RNA (tRNA) modifications and modifying enzymes of the coconut in this survey. An extraction method for the tRNA pool from coconut flesh was built. In total, 33 species of modified nucleosides and 66 homologous genes of modifying enzymes were confirmed using a nucleoside analysis using high-performance liquid chromatography combined with high-resolution mass spectrometry (HPLC-HRMS) and homologous protein sequence alignment. The positions of tRNA modifications, including pseudouridines, were preliminarily mapped using a oligonucleotide analysis, and the features of their modifying enzymes were summarized. Interestingly, we found that the gene encoding the modifying enzyme of 2'-O-ribosyladenosine at the 64th position of tRNA (Ar(p)64) was uniquely overexpressed under high-salinity stress. In contrast, most other tRNA-modifying enzymes were downregulated with mining transcriptomic sequencing data. According to previous physiological studies of Ar(p)64, the coconut appears to enhance the quality control of the translation process when subjected to high-salinity stress. We hope this survey can help advance research on tRNA modification and scientific studies of the coconut, as well as thinking of the safety and nutritional value of naturally modified nucleosides.

Downregulation of miR-146a-5p Promotes Acute Pancreatitis through Activating the TLR9/NLRP3 Signaling Pathway by Targeting TRAF6 In Vitro Rat Model.

Acute pancreatitis (AP) is mainly caused by acinar cells releasing various inflammatory factors, causing inflammatory storms and leading to severe pancreatitis. Detection methods and treatment targets for pancreatitis are lacking, raising the urgency of identifying diagnostic markers and therapeutic targets for AP. MicroRNAs (miRNAs) have recently been identified as molecular markers for various biological processes such as tumors, immunity, and metabolism, and the involvement of miRNAs in inflammatory responses has been increasingly studied. To explore the role of miRNAs in AP is the primary objective of this study. By using qPCR on our cerulein-induced pancreatitis cell model, it is worth noting that the change of miR-146a-5p expression in inflammation-related miRNAs in AP was predominant. Next, ELISA, CCK8, and flow cytometry were used to inspect the impact of miR-146a-5p on pancreatitis. BiBiServ bioinformatics anticipated binding ability of miR-146a-5p and 3'-untranslated region (3'UTR) of TNF receptor-associated factor 6 (TRAF6), and the dual-luciferase assay verified the combination of the two. TRAF6 knockdown verified the effect of TRAF6 on the progression of pancreatitis. Finally, rescue experiments verified the capability of miR-146a-5p and TRAF6 interaction on the Toll-like receptor 9 (TLR9)/NOD-like receptor protein 3 (NLRP3) signaling pathway and cell function. The expression of miR-146a-5p decreased in cerulein-induced AR42J pancreatic acinar cells. Functional experiments verified that miR-146a-5p facilitated the proliferation of AR42J pancreatic acinar cells and inhibited their apoptosis. Bioinformatic predictions and dual-luciferase experiments verified the actual binding efficiency between miR-146a-5p and 3'UTR of TRAF6. Our study confirmed that knockdown of TRAF6 restrained the progression of pancreatitis, and knockdown of TRAF6 rescued pancreatitis caused by miR-146a-5p downregulation by the TLR9/NLRP3 signaling pathway. Therefore, downregulation of miR-146a-5p in the induced pancreatitis cell model promotes the progression of pancreatitis via the TLR9/TRAF6/NLRP3 signaling pathway. There is potential for miR-146a-5p to serve as a diagnostic marker and therapeutic nucleic acid drug for AP.

Activation of the Nuclear Factor-kappa B Signaling Pathway Damages the Epithelial Barrier in the Human Pancreatic Ductal Adenocarcinoma Cell Line HPAF-II.

OBJECTIVES: Injury of the pancreatic duct epithelial barrier plays a critical role in the development of acute pancreatitis. The activity of the nuclear factor-kappa B (NF-κB) pathway is involved in the disruption of the pancreatic duct epithelial barrier. This study investigated how NF-κB impacts the dysfunction of the pancreatic duct epithelial barrier. METHODS: A human pancreatic ductal adenocarcinoma cell line was treated with tumor necrosis factor-alpha (TNF-α) and pyrrolidine dithiocarbamate. The expression levels of p65 and p-p65 were detected to evaluate NF-κB activity. Tricellulin (TRIC) expression levels were measured to assess the change in tight junction (TJ)-related proteins. The expression and localization of myosin light chain kinase (MLCK) were investigated. The structure of TJs and monolayer permeability were also examined. RESULTS: NF-κB was activated by TNF-α and suppressed by pyrrolidine dithiocarbamate. Activation of NF-κB upregulated the expression levels of TRIC and MLCK. Broadened TJs were observed after NF-κB was activated. Lower monolayer permeability was observed when NF-κB was suppressed. CONCLUSIONS: Activation of the NF-κB pathway induced by TNF-α leads to increased TRIC and MLCK expression, resulting in broadened TJs and high permeability, which contribute to damage to the pancreatic duct epithelial barrier.