Prevalence of antibiotic-resistant Acinetobacter spp. on soil and crops collected from agricultural fields in South Korea.

The emergence of antibiotic resistance in Acinetobacter spp. is a rising public health concern worldwide. The objective of this study was to investigate the prevalence of antibiotic-resistance genes and the virulence of Acinetobacter spp. isolated from soil and crops obtained from agricultural fields in South Korea. Eight Acinetobacter spp. isolates carried various antibiotic resistance genes, such as emrAB (100%), cat/craA (100%), and aadA gene (87.5%). Minimum inhibitory concentration (MIC) analysis revealed that strains harboring antibiotic resistance genes exhibited high resistance to the respective antibiotics, such as colistin, chloramphenicol, and streptomycin. Interestingly, most of these isolates had high capability of biofilm formation and swarming motility, along with faster growth rates. Taken together, our study demonstrated that antibiotic-resistant Acinetobacter isolated from agricultural settings in South Korea not only frequently carries antibiotic resistance genes but also has virulence-related traits. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-023-01496-7.

Complete genome sequence of Pectobacterium brasiliense strain 21PCA_AGRO2 with antimicrobial resistance isolated from napa cabbage.

We report a complete genome of Pectobacterium brasiliense strain 21PCA_AGRO2 isolated from napa cabbage, in which the genome consists of a circular chromosome comprising 4,919,671 bp with 4,399 coding DNA sequences, 22 rRNA genes, 77 tRNA genes, and 9 noncoding RNA genes.

HGTree v2.0: a comprehensive database update for horizontal gene transfer (HGT) events detected by the tree-reconciliation method.

HGTree is a database that provides horizontal gene transfer (HGT) event information on 2472 prokaryote genomes using the tree-reconciliation method. HGTree was constructed in 2015, and a large number of prokaryotic genomes have been additionally published since then. To cope with the rapid rise of prokaryotic genome data, we present HGTree v2.0 (http://hgtree2.snu.ac.kr), a newly updated version of our HGT database with much more extensive data, including a total of 20 536 completely sequenced non-redundant prokaryotic genomes, and more reliable HGT information results curated with various steps. As a result, HGTree v2.0 has a set of expanded data results of 6 361 199 putative horizontally transferred genes integrated with additional functional information such as the KEGG pathway, virulence factors and antimicrobial resistance. Furthermore, various visualization tools in the HGTree v2.0 database website provide intuitive biological insights, allowing the users to investigate their genomes of interest.

The RNA-binding protein, HuD regulates proglucagon biosynthesis in pancreatic α cells.

Glucagon is a peptide hormone generated by pancreatic α cells. It is the counterpart of insulin and plays an essential role in the regulation of blood glucose level. Therefore, a tight regulation of glucagon levels is pivotal to maintain homeostasis of blood glucose. However, little is known about the mechanisms regulating glucagon biosynthesis. In this study, we demonstrate that the RNA-binding protein HuD regulates glucagon expression in pancreatic α cells. HuD was found in α cells from mouse pancreatic islet and mouse glucagonoma αTC1 cell line. Ribonucleoprotein immunoprecipitation analysis, followed by RT-qPCR showed the association of HuD with glucagon mRNA. Knockdown of HuD resulted in a reduction in both proglucagon expression and cellular glucagon level by decreasing its de novo synthesis. Reporter analysis using the EGFP reporter containing 3' untranslated region (3'UTR) of glucagon mRNA showed that HuD regulates proglucagon expression via its 3'UTR. In addition, the relative level of glucagon in the islets and plasma was lower in HuD knockout (KO) mice compared to age-matched control mice. Taken together, these results suggest that HuD is a novel factor regulating the biosynthesis of proglucagon in pancreatic α cells.

RNA binding protein HuD and microRNA-203a cooperatively regulate insulinoma-associated 1 mRNA.

RNA binding protein HuD regulates translation and turnover of target mRNAs, thereby affecting gene expression at the posttranscriptional level in mainly neuronal as well as pancreatic ß-cells. Here, we identified insulinoma-associated 1 (INSM1), an essential factor governing differentiation and proliferation of neuroendocrine cells, as a novel target of HuD and demonstrated the regulatory mechanism of INSM1 expression by HuD. HuD bound to 3'untranslated region (3'UTR) of Insm1 mRNA and negatively regulated its expression; knockdown of HuD increased INSM1 expression, while HuD overexpression repressed it by destabilizing its mRNA. In addition, we further demonstrated that HuD enhanced reduction of INSM1 by miR-203a, a novel miRNA targeting Insm1 mRNA 3'UTR. These results suggest that HuD and miR-203a cooperatively regulate INSM1 expression and it provides a novel regulatory mechanism of INSM1 expression by HuD and miR-203a.

RNA binding protein HuD contributes to ß-cell dysfunction by impairing mitochondria dynamics.

Imbalanced mitochondrial dynamics in pancreatic ß-cells contributes to ß-cell dysfunction in diabetes; however, the molecular mechanisms underlying mitochondrial dynamics in the pathology of diabetes are not fully elucidated. We previously reported the reduction of RNA binding protein HuD in pancreatic ß-cells of diabetes. Herein, we demonstrate that HuD plays a novel role in the regulation of mitochondrial dynamics by promoting mitochondrial fusion. We show enhanced mitochondrial fragmentation in the pancreas of db/db mice and HuD KO mice. Downregulation of HuD increases the number of cells with fragmented mitochondria and reduces the mitochondrial activity determined by mitochondrial membrane potential and ATP production in mouse insulinoma ßTC6 cells. HuD binds to 3'-untraslated region of mitofusin 2 (Mfn2) mRNA and positively regulates its expression. Ectopic expression of Mfn2 in ßTC6 cells stably expressing short hairpin RNA against HuD (shHuD) restores HuD-mediated mitochondrial dysfunction. Taken together, our results suggest that HuD regulates mitochondrial dynamics by regulating Mfn2 level and its reduced expression leads to mitochondrial dysfunction in pancreatic ß-cells.

Gas Separation Silica Membranes Prepared by Chemical Vapor Deposition of Methyl-Substituted Silanes.

The effect on the gas permeance properties and structural morphology of the presence of methyl functional groups in a silica membrane was studied. Membranes were synthesized via chemical vapor deposition (CVD) at 650 °C and atmospheric pressure using three silicon compounds with differing numbers of methyl- and methoxy-functional groups: tetramethyl orthosilicate (TMOS), methyltrimethoxysilane (MTMOS), and dimethyldimethoxysilane (DMDMOS). The residence time of the silica precursors in the CVD process was adjusted for each precursor and optimized in terms of gas permeance and ideal gas selectivity criteria. Final H2 permeances at 600 °C for the TMOS-, MTMOS-, and DMDMOS-derived membranes were respectively 1.7 × 10-7, 2.4 × 10-7, and 4.4 × 10-8 mol∙m-2∙s-1∙Pa-1 and H2/N2 selectivities were 990, 740, and 410. The presence of methyl groups in the membranes fabricated with the MTMOS and DMDMOS precursors was confirmed via Fourier-transform infrared (FTIR) spectroscopy. From FTIR analysis, an increasing methyl signal in the silica structure was correlated with both an improvement in the hydrothermal stability and an increase in the apparent activation energy for hydrogen permeation. In addition, the permeation mechanism for several gas species (He, H2, Ne, CO2, N2, and CH4) was determined by fitting the gas permeance temperature dependence to one of three models: solid state, gas-translational, or surface diffusion.

Fabrication and Evaluation of Trimethylmethoxysilane (TMMOS)-Derived Membranes for Gas Separation.

Gas separation membranes were fabricated with varying trimethylmethoxysilane(TMMOS)/tetraethoxy orthosilicate (TEOS) ratios by a chemical vapor deposition (CVD) method at650 °C and atmospheric pressure. The membrane had a high H2 permeance of 8.3 × 10-7 mol m-2 s-1Pa-1 with H2/CH4 selectivity of 140 and H2/C2H6 selectivity of 180 at 300 °C. Fourier transforminfrared (FTIR) measurements indicated existence of methyl groups at high preparationtemperature (650 °C), which led to a higher hydrothermal stability of the TMMOS-derivedmembranes than of a pure TEOS-derived membrane. Temperature-dependence measurements ofthe permeance of various gas species were used to establish a permeation mechanism. It was foundthat smaller species (He, H2, and Ne) followed a solid-state diffusion model while larger species (N2,CO2, and CH4) followed a gas translational diffusion model.

A miR-194/PTBP1/CCND3 axis regulates tumor growth in human hepatocellular carcinoma.

Polypyrimidine tract-binding protein 1 (PTBP1) is one of the most investigated multifunctional RNA-binding proteins (RBP), controlling almost all steps of mRNA metabolism and processing. It has been reported that PTBP1 is overexpressed in many different types of cancer and this high expression is associated with increased proliferation and poor prognoses. However, there are no reports on a putative role for PTBP1 in the molecular abnormalities and pathogenesis of hepatocellular carcinoma (HCC). Here, we identified PTBP1 as a positive regulator of human HCC growth. The expression of PTBP1 was increased in human HCC cells and tissues compared to the corresponding controls, and this high expression was positively correlated with increased tumor size and a reduced survival rate. Mechanistically, PTBP1 enhanced cyclin D3 (CCND3) translation by interacting with the 5'-untranslated region (5'-UTR) of CCND3 mRNA, consequently facilitating cell cycle progression and tumor growth. Furthermore, we found that miR-194 inhibits PTBP1 expression by binding to the 3'-UTR of PTBP1 mRNA, resulting in reduced CCND3 levels and HCC cell growth; moreover, the levels of PTBP1 were negatively correlated with miR-194 levels in HCC. Taken together, these findings identify PTBP1 as a pivotal enhancer of HCC growth; the miR-194/PTBP1/CCND3 axis seemingly has a crucial role in the development and progression of HCC and targeting the axis could be a novel therapeutic strategy against human HCC. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Heterogeneous nuclear ribonucleoprotein A1 promotes the expression of autophagy-related protein 6 in human colorectal cancer.

Autophagy, a lysosomal self-degradative process of cellular components, is essential for cellular homeostasis to response cellular stress and is tightly controlled by autophagy-related genes (ATGs). Autophagy-related gene 6 (ATG6, also known as Beclin-1 in human) is an essential factor regulating autophagy and apoptosis. RNA binding proteins (RBPs) regulate gene expression at the post-transcriptional level and their differential expression is linked to the pathogenesis of several human diseases. Here, we demonstrate the role of heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1) as a novel factor regulating ATG6 expression. hnRNPA1 associates with the 3' untranslated region (3'UTR) of ATG6 mRNA and promotes its expression without significant changes at the mRNA level. Knockdown of hnRNPA1 decreases ATG6 expression, which is enhanced by the overexpression of hnRNPA1. Also, we show augmented expression of both hnRNPA1 and ATG6 in the colorectal cancer (CRC) tissues obtained from patients and demonstrate a positive correlation of their expression in CRC tissues. Our results suggest the potential role of hnRNPA1-mediated ATG6 regulation in the pathogenesis of CRC.

The MicroRNA-551a/MEF2C Axis Regulates the Survival and Sphere Formation of Cancer Cells in Response to 5-Fluorouracil.

microRNAs regulate a diverse spectrum of cancer biology, including tumorigenesis, metastasis, stemness, and drug resistance. To investigate miRNA-mediated regulation of drug resistance, we characterized the resistant cell lines to 5-fluorouracil by inducing stable expression of miRNAs using lenti-miRNA library. Here, we demonstrate miR-551a as a novel factor regulating cell survival after 5-FU treatment. miR-551a-expressing cells (Hep3B-lenti-miR-551a) were resistant to 5-FU-induced cell death, and after 5-FU treatment, and showed significant increases in cell viability, cell survival, and sphere formation. It was further shown that myocyte-specific factor 2C is the direct target of miR-551a. Our results suggest that miR-551a plays a novel function in regulating 5-FU-induced cell death, and targeting miR-551a might be helpful to sensitize cells to anti-cancer drugs.

RNA Binding Protein HuR Promotes Autophagosome Formation by Regulating Expression of Autophagy-Related Proteins 5, 12, and 16 in Human Hepatocellular Carcinoma Cells.

Autophagy is a process of lysosomal self-degradation of cellular components by forming autophagosomes. Autophagosome formation is an essential process in autophagy and is fine-tuned by various autophagy-related gene (ATG) products, including ATG5, ATG12, and ATG16. Although several reports have shown that numerous factors affect multiple levels of gene regulation to orchestrate cellular autophagy, the detailed mechanism of autophagosome formation still needs further investigation. In this study, we demonstrate that the RNA binding protein HuR (human antigen R) performs an essential function in autophagosome formation. We observe that HuR silencing leads to inhibition of autophagosome formation and autophagic flux in liver cells. Ribonucleoprotein immunoprecipitation (RIP) assay allows the identification of ATG5, ATG12, and ATG16 mRNAs as the direct targets of HuR. We further show that HuR mediates the translation of ATG5, ATG12, and ATG16 mRNAs by binding to their 3' untranslated regions (UTRs). In addition, we show that HuR expression positively correlates with the levels of ATG5 and ATG12 in hepatocellular carcinoma (HCC) cells. Collectively, our results suggest that HuR functions as a pivotal regulator of autophagosome formation by enhancing the translation of ATG5, ATG12, and ATG16 mRNAs and that augmented expression of HuR and ATGs may participate in the malfunction of autophagy in HCC cells.

Reduced expression of the RNA-binding protein HuD in pancreatic neuroendocrine tumors correlates with low p27Kip1 levels and poor prognosis.

For the majority of patients diagnosed with pancreatic neuroendocrine tumors (NETs), there is significant malignant potential with a poor prognosis; however, the molecular abnormalities and pathogenesis of pancreatic NETs have not been firmly established. Here, we report that loss of expression of the RNA-binding protein HuD correlates with low p27Kip1 (p27) levels and poor prognosis in pancreatic NETs. HuD expression was frequently lost in many human pancreatic NETs, and these pancreatic NETs showed aggressive clinicopathological phenotypes with low p27 levels, increased tumor size, higher World Health Organization grade and pT stage of the tumor, and the presence of angioinvasion. Furthermore, loss of HuD was an independent, progression-free prognostic factor in multivariate survival analysis. However, the level of HuR, a member of the same Hu protein family as HuD, was not significantly correlated with pancreatic NET size and progression. Mechanistically, HuD enhanced p27 mRNA translation by interacting with both the 5'-untranslated region (UTR) and the 3'-UTR of p27 mRNA, and consequently suppressed cell cycle progression and tumor growth. In addition, HuD competed with miR-30a-3p for binding to the 3'-UTR of p27 mRNA, suggesting an interplay between HuD and miR-30a-3p in controlling p27 translation. Our results identify HuD as a pivotal suppressor of pancreatic NET growth, and suggest that HuD has potential value as a prognostic factor of pancreatic NETs. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Comparative Genomic Analysis of Lactobacillus plantarum GB-LP1 Isolated from Traditional Korean Fermented Food.

As probiotics play an important role in maintaining a healthy gut flora environment through antitoxin activity and inhibition of pathogen colonization, they have been of interest to the medical research community for quite some time now. Probiotic bacteria such as Lactobacillus plantarum, which can be found in fermented food, are of particular interest given their easy accessibility. We performed whole-genome sequencing and genomic analysis on a GB-LP1 strain of L. plantarum isolated from Korean traditional fermented food; this strain is well known for its functions in immune response, suppression of pathogen growth, and antitoxin effects. The complete genome sequence of GB-LP1 is a single chromosome of 3,040,388 bp with 2,899 predicted open reading frames. Genomic analysis of GB-LP1 revealed two CRISPR regions and genes showing accelerated evolution, which may have antibiotic and antitoxin functions. The aim of the present study was to predict strain specific-genomic characteristics and assess the potential of this new strain as lactic acid bacteria at the genomic level using in silico analysis. These results provide insight into the L. plantarum species as well as confirm the possibility of its utility as a candidate probiotic.

microRNA-200a-3p increases 5-fluorouracil resistance by regulating dual specificity phosphatase 6 expression.

Acquisition of resistance to anti-cancer drugs is a significant obstacle to effective cancer treatment. Although several efforts have been made to overcome drug resistance in cancer cells, the detailed mechanisms have not been fully elucidated. Here, we investigated whether microRNAs (miRNAs) function as pivotal regulators in the acquisition of anti-cancer drug resistance to 5-fluorouracil (5-FU). A survey using a lentivirus library containing 572 precursor miRNAs revealed that five miRNAs promoted cell survival after 5-FU treatment in human hepatocellular carcinoma Hep3B cells. Among the five different clones, the clone expressing miR-200a-3p (Hep3B-miR-200a-3p) was further characterized as a 5-FU-resistant cell line. The cell viability and growth rate of Hep3B-miR-200a-3p cells were higher than those of control cells after 5-FU treatment. Ectopic expression of a miR-200a-3p mimic increased, while inhibition of miR-200a-3p downregulated, cell viability in response to 5-FU, doxorubicin, and CDDP (cisplatin). We also showed that dual-specificity phosphatase 6 (DUSP6) is a novel target of miR-200a-3p and regulates resistance to 5-FU. Ectopic expression of DUSP6 mitigated the pro-survival effects of miR-200a-3p. Taken together, these results lead us to propose that miR-200a-3p enhances anti-cancer drug resistance by decreasing DUSP6 expression.

Comparative genomic analysis reveals genetic features related to the virulence of Bacillus cereus FORC_013.

BACKGROUND: Bacillus cereus is well known as a gastrointestinal pathogen that causes food-borne illness. In the present study, we sequenced the complete genome of B. cereus FORC_013 isolated from fried eel in South Korea. To extend our understanding of the genomic characteristics of FORC_013, we conducted a comparative analysis with the published genomes of other B. cereus strains. RESULTS: We fully assembled the single circular chromosome (5,418,913 bp) and one plasmid (259,749 bp); 5511 open reading frames (ORFs) and 283 ORFs were predicted for the chromosome and plasmid, respectively. Moreover, we detected that the enterotoxin (NHE, HBL, CytK) induces food-borne illness with diarrheal symptom, and that the pleiotropic regulator, along with other virulence factors, plays a role in surviving and biofilm formation. Through comparative analysis using the complete genome sequence of B. cereus FORC_013, we identified both positively selected genes related to virulence regulation and 224 strain-specific genes of FORC_013. CONCLUSIONS: Through genome analysis of B. cereus FORC_013, we identified multiple virulence factors that may contribute to pathogenicity. These results will provide insight into further studies regarding B. cereus pathogenesis mechanism at the genomic level.

The miR-24-3p/p130Cas: a novel axis regulating the migration and invasion of cancer cells.

MicroRNAs (miRNAs) are small non-coding RNAs that negatively regulate gene expression by suppressing translation or facilitating mRNA decay. Differential expression of miRNAs is involved in the pathogenesis of several diseases including cancer. Here, we investigated the role of-miR-24-3p as a downregulated miRNA in metastatic cancer. miR-24-3p was decreased in metastatic cancer and lower expression of miR-24-3p was related to poor survival of cancer patients. Consistently, ectopic expression of miR-24-3p suppressed the cell migration, invasion, and proliferation of MCF7, Hep3B, B16F10, SK-Hep1, and PC-3 cells by directly targeting p130Cas. Stable expression of p130Cas restored miR-24-3p-mediated inhibition of cell migration and invasion. These results suggest that miR-24-3p functions as a tumor suppressor and the miR-24-3p/p130Cas axis is a novel factor of cancer progression by regulating cell migration and invasion.

Detecting Positive Selection of Korean Native Goat Populations Using Next-Generation Sequencing.

Goats (Capra hircus) are one of the oldest species of domesticated animals. Native Korean goats are a particularly interesting group, as they are indigenous to the area and were raised in the Korean peninsula almost 2,000 years ago. Although they have a small body size and produce low volumes of milk and meat, they are quite resistant to lumbar paralysis. Our study aimed to reveal the distinct genetic features and patterns of selection in native Korean goats by comparing the genomes of native Korean goat and crossbred goat populations. We sequenced the whole genome of 15 native Korean goats and 11 crossbred goats using next-generation sequencing (Illumina platform) to compare the genomes of the two populations. We found decreased nucleotide diversity in the native Korean goats compared to the crossbred goats. Genetic structural analysis demonstrated that the native Korean goat and crossbred goat populations shared a common ancestry, but were clearly distinct. Finally, to reveal the native Korean goat's selective sweep region, selective sweep signals were identified in the native Korean goat genome using cross-population extended haplotype homozygosity (XP-EHH) and a cross-population composite likelihood ratio test (XP-CLR). As a result, we were able to identify candidate genes for recent selection, such as the CCR3 gene, which is related to lumbar paralysis resistance. Combined with future studies and recent goat genome information, this study will contribute to a thorough understanding of the native Korean goat genome.

Complete genome of Vibrio parahaemolyticus FORC014 isolated from the toothfish.

BACKGROUND: Foodborne illness can occur due to various pathogenic bacteria such as Staphylococcus aureus, Escherichia coli and Vibrio parahaemolyticus, and can cause severe gastroenteritis symptoms. In this study, we completed the genome sequence of a foodborne pathogen V. parahaemolyticus FORC_014, which was isolated from suspected contaminated toothfish from South Korea. Additionally, we extended our knowledge of genomic characteristics of the FORC_014 strain through comparative analysis using the complete sequences of other V. parahaemolyticus strains whose complete genomes have previously been reported. RESULTS: The complete genome sequence of V. parahaemolyticus FORC_014 was generated using the PacBio RS platform with single molecule, real-time (SMRT) sequencing. The FORC_014 strain consists of two circular chromosomes (3,241,330 bp for chromosome 1 and 1,997,247 bp for chromosome 2), one plasmid (51,383 bp), and one putative phage sequence (96,896 bp). The genome contains a total of 4274 putative protein coding sequences, 126 tRNA genes and 34 rRNA genes. Furthermore, we found 33 type III secretion system 1 (T3SS1) related proteins and 15 type III secretion system 2 (T3SS2) related proteins on chromosome 1. This is the first reported result of Type III secretion system 2 located on chromosome 1 of V. parahaemolyticus without thermostable direct hemolysin (tdh) and thermostable direct hemolysin-related hemolysin (trh). CONCLUSIONS: Through investigation of the complete genome sequence of V. parahaemolyticus FORC_014, which differs from previously reported strains, we revealed two type III secretion systems (T3SS1, T3SS2) located on chromosome 1 which do not include tdh and trh genes. We also identified several virulence factors carried by our strain, including iron uptake system, hemolysin and secretion system. This result suggests that the FORC_014 strain may be one pathogen responsible for foodborne illness outbreak. Our results provide significant genomic clues which will assist in future understanding of virulence at the genomic level and help distinguish between clinical and non-clinical isolates.