Crude Oil Contamination Enhances the Lipoxygenase Gene Expression in the Green Microalga Scenedesmus dimorphu

Crude oil is a mixture of hydrocarbons, which are mainly toxic to living organisms. Over recent years it has become clear that algae may play a substantial role in the biodegradation of the crude oil harmful hydrocarbons by their dioxygenase system. The present study was conducted to evaluate alterations in the expression levels of lipoxygenase gene (lox) in Scenedesmus dimorphus after exposure to crude oil. The extraction of total RNA was performed by RNX plus reagent. After cDNA synthesis, a qRT-PCR analysis was performed to determine the expression level of the lox gene. The acquired data were analyzed by SPSS Statistics. Based on the obtained results, crude oil treatments (0.02 and 1%) led to a relatively high expression level of lox compared to control conditions. Moreover, incubation time significantly affected the relative expression of this gene in both oil-treated and non-treated algae. The outcomes of this research indicated that S. dimorphus has a high potential for the enhancement of enzymatic activity under crude oil treatment through increased lox expression.

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Investigation of olfactory receptor family 51 subfamily j member 1 (OR51J1) gene susceptibility as a potential breast cancer-associated biomarker

PLoS ONE ◽

10.1371/journal.pone.0246752 ◽

2021 ◽

Vol 16 (2) ◽

pp. e0246752

Author(s):

Maryam Asadi ◽

Nahid Ahmadi ◽

Simin Ahmadvand ◽

Ali Akbar Jafari ◽

Akbar Safaei ◽

...

Keyword(s):

Breast Cancer ◽

Statistical Analysis ◽

Protein Expression ◽

Olfactory Receptor ◽

Mrna Level ◽

Expression Level ◽

Pcr Analysis ◽

Normal Tissues ◽

Qrt Pcr ◽

Human Protein Atlas

Among cancer treatment methods, targeted therapy using cancer-associated biomarkers has minimum side effects. Recently olfactory receptor (OR) family attracts the researcher’s attention as a favorable biomarker of cancer. Here, a statistical approach using complete data from the human protein atlas database was used to evaluate the potential of OR51J1 gene as a cancer-associated biomarker. To confirm the findings of statistical analysis, the OR51J1 mRNA and protein expression levels in breast tumor and normal tissue were measured using quantitative Real Time PCR (qRT-PCR) and immunohistochemistry (IHC) techniques. The association with clinicopathological factors was analyzed. Statistical analysis revealed that OR51J1 has a high expression level in more than 20 types of cancer tissues without any expression in 44 normal tissues. In 15 cancer types, including breast cancer, expression score was more than 90%. The qRT-PCR analysis in breast cancer showed OR51J1 have significantly higher expression level in tumors than normal tissues (2.91 fold). The IHC results showed OR51J1 expression on other cellular subtypes than tumor and normal cells, including myoepithelium, fibroblast, and lymphocytes. OR51J1 protein expression in invasive cells, as well as its overall score, showed a significant correlation with ER and PR expression and breast cancer (BC) subtypes. Results revealed the potential of OR51J1 as a cancer-associated biomarker for the diagnosis of breast cancer at the mRNA level.

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Abstract 16892: Beneficial Effect of miR-25 Decoy Overexpression in a Murine Model of Heart Failure

Circulation ◽

10.1161/circ.130.suppl_2.16892 ◽

2014 ◽

Vol 130 (suppl_2) ◽

Author(s):

Dongtak Jeong ◽

Jae Gyun Oh ◽

Alessia Baccarini ◽

Brian Brown ◽

Mark Mercola ◽

...

Keyword(s):

Heart Failure ◽

Cardiac Function ◽

Therapeutic Potential ◽

Cardiac Contractility ◽

Pressure Overload ◽

Fold Increase ◽

Expression Level ◽

Pcr Analysis ◽

Qrt Pcr ◽

Adult Cardiomyocytes

Introduction: Recently, our group found that miR-25 is a key microRNA that regulates SERCA2a and we showed anti-miR25 treatment enhanced cardiac contractility and function through SERCA2a restoration in murine heart failure model. However, the strong and stable suppression of specific microRNA activity would be essential to evaluate the therapeutic potential of such an approach. In this study, we constructed a miR-25 decoy t using TuD RNA (tough decoy RNAs) and miR-25 decoy activity was evaluated in various settings. Methods: First, miR-25 decoy construct was applied to both isolated adult cardiomyocytes and H9C2 cardiomyoblasts. A pMirTarget vector containing SERCA2a 3’-UTR under luciferase was used to evaluate this decoy specificity. Second, we generated pressure-overload heart failure models in mouse by TAC operation. In the HF mice, AAV9-GFP (control) and AAV9-miR-25 decoy were delivered by tail vein injection. Two months following gene transfer, cardiac function was evaluated by echocardiography and invasive hemodynamics. Protein and RNA expression levels of SERCA2a and miR-25 expression level were confirmed by qRT-PCR analysis. Results: First, we observed that we were able to achieve about 2-3 fold increase of SERCA2a expression by miR-25 decoy transfection in both H9C2 and in isolated adult cardiomyocytes. Also, similar expression pattern was confirmed in the heart of miR-25 decoy injected normal mice. Second, the HF model by TAC surgery was confirmed with echocardiography. Overall average FS (%) in HF was 37.77+/- 8.75 (n=10) and in non-surgery control mice was 62.51 +/- 3.42(n=4). After AAV9 injection, cardiac function of AAV9-miR-25 decoy injected mice was enhanced but AAV9-GFP injected mice showed severe cardiac dysfunction and dilation (AAV-GFP (n=6) vs AAV-miR-25 decoy (n=4)). Third, western blot analysis showed that SERCA2a expression was significantly restored in miR-25 decoy injected mice. In addition, we confirmed that miR-25 expression level was kept low by qRT-PCR analysis. Taken together, our data would indicate that using miR-25 decoy is an effective strategy for the long-term suppression of miR-25 and it may be a promising therapeutic target to restore SERCA2a and reverse the HF phenotype.

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Cinnamaldehyde inhibits the growth of Phytophthora capsici through disturbing metabolic homoeostasis

PeerJ ◽

10.7717/peerj.11339 ◽

2021 ◽

Vol 9 ◽

pp. e11339

Author(s):

Yinan Wang ◽

Mengke Wang ◽

Min Li ◽

Te Zhao ◽

Lin Zhou

Keyword(s):

Fatty Acid ◽

Protein Kinase ◽

Fatty Acid Metabolism ◽

Acid Metabolism ◽

Phytophthora Capsici ◽

Enrichment Analysis ◽

Functional Enrichment ◽

Expression Level ◽

Pcr Analysis ◽

Qrt Pcr

Background Phytophthora capsici Leonian (P. capsici) can cause wilting and roots rotting on pepper and other cash crops. The new fungicide cinnamaldehyde (CA) has high activity against this pathogen. However, its potential mechanism is still unknown. Methods In order to gain insights into the mechanism, isobaric tags for relative and absolute quantification (iTRAQ)-based quantitative proteomics was used to analyze P. capsici treated with CA. The iTRAQ results were evaluated by parallel reaction monitoring (PRM) analysis and quantitative real-time PCR (qRT-PCR) analysis. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was used to speculate the biochemical pathways that the agent may act on. Results The results showed that 1502 differentially expressed proteins were identified, annotated and classified into 209 different terms (like metabolic process, cellular process, single-organism process) based on Gene Ontology (GO) functional enrichment analysis and nine different pathways (glyoxylate and dicarboxylate metabolism, fatty acid metabolism and so on) based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. This study suggested that CA disordered fatty acid metabolism, polysaccharide metabolism and leucine metabolism. Based on PRM analysis, five proteins including CAMK/CAMK1 protein kinase, glucan 1,3-beta-glucosidase, 1,3-beta-glucanosyltransferase, methylcrotonoyl-CoA carboxylase subunit alpha and isovaleryl-CoA dehydrogenase were down-regulated in P. capsici treated with CA. Furthermore, the qRT-PCR analysis showed that the gene expression level of the interested proteins was consistent with the protein expression level, except for CAMK/CAMK1 protein kinase, acetyl-CoA carboxylase and fatty acid synthase subunit alpha. Conclusions CA destroyed the metabolic homoeostasisof P. capsici, which led to cell death. This is the first proteomic analysis of P. capsici treated with CA, which may provide an important information for exploring the mechanism of the fungicide CA against P. capsici.

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ITRAQ-based quantitative proteomic analysis of Fusarium moniliforme (Fusarium verticillioides) in response to Phloridzin inducers

Proteome Science ◽

10.1186/s12953-021-00170-2 ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Rong Zhang ◽

Weitao Jiang ◽

Xin Liu ◽

Yanan Duan ◽

Li Xiang ◽

...

Keyword(s):

Fusarium Moniliforme ◽

Abiotic Factors ◽

Fusarium Verticillioides ◽

Protein Profile ◽

Sugar Metabolism ◽

Differentially Expressed ◽

Pcr Analysis ◽

Differentially Expressed Proteins ◽

Apple Replant Disease ◽

Qrt Pcr

Abstract Background Apple replant disease (ARD) has been reported from all major fruit-growing regions of the world, and is often caused by biotic factors (pathogen fungi) and abiotic factors (phenolic compounds). In order to clarify the proteomic differences of Fusarium moniliforme under the action of phloridzin, and to explore the potential mechanism of F. moniliforme as the pathogen of ARD, the role of Fusarium spp. in ARD was further clarified. Methods In this paper, the quantitative proteomics method iTRAQ analysis technology was used to analyze the proteomic differences of F. moniliforme before and after phloridzin treatment. The differentially expressed protein was validated by qRT-PCR analysis. Results A total of 4535 proteins were detected, and 293 proteins were found with more than 1.2 times (P< 0.05) differences. In-depth data analysis revealed that 59 proteins were found with more than 1.5 times (P< 0.05) differences, and most proteins were consistent with the result of qRT-PCR. Differentially expressed proteins were influenced a variety of cellular processes, particularly metabolic processes. Among these metabolic pathways, a total of 8 significantly enriched KEGG pathways were identified with at least 2 affiliated proteins with different abundance in conidia and mycelium. Functional pathway analysis indicated that up-regulated proteins were mainly distributed in amino sugar, nucleotide sugar metabolism, glycolysis/ gluconeogenesis and phagosome pathways. Conclusions This study is the first to perform quantitative proteomic investigation by iTRAQ labeling and LC-MS/MS to identify differentially expressed proteins in F. moniliforme under phloridzin conditions. The results confirmed that F. moniliforme presented a unique protein profile that indicated the adaptive mechanisms of this species to phloridzin environments. The results deepened our understanding of the proteome in F. moniliforme in response to phloridzin inducers and provide a basis for further exploration for improving the efficiency of the fungi as biocontrol agents to control ARD.

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Downregulation of lncRNA ZNF582-AS1 due to DNA hypermethylation promotes clear cell renal cell carcinoma growth and metastasis by regulating the N(6)-methyladenosine modification of MT-RNR1

Journal of Experimental & Clinical Cancer Research ◽

10.1186/s13046-021-01889-8 ◽

2021 ◽

Vol 40 (1) ◽

Author(s):

Wuping Yang ◽

Kenan Zhang ◽

Lei Li ◽

Yawei Xu ◽

Kaifang Ma ◽

...

Keyword(s):

Dna Methylation ◽

Renal Cell Carcinoma ◽

Protein Expression ◽

Cell Carcinoma ◽

Clinical Significance ◽

Renal Cell ◽

Clear Cell ◽

Expression Level ◽

Cell Renal Cell Carcinoma ◽

Qrt Pcr

Abstract Background Emerging evidence confirms that lncRNAs (long non-coding RNAs) are potential biomarkers that play vital roles in tumors. ZNF582-AS1 is a novel lncRNA that serves as a potential prognostic marker of cancers. However, the specific clinical significance and molecular mechanism of ZNF582-AS1 in ccRCC (clear cell renal cell carcinoma) are unclear. Methods Expression level and clinical significance of ZNF582-AS1 were determined by TCGA-KIRC data and qRT-PCR results of 62 ccRCCs. DNA methylation status of ZNF582-AS1 promoter was examined by MSP, MassARRAY methylation and demethylation analysis. Gain-of-function experiments were conducted to investigate the biological roles of ZNF582-AS1 in the phenotype of ccRCC. The subcellular localization of ZNF582-AS1 was detected by RNA FISH. iTRAQ, RNA pull-down and RIP-qRT-PCR were used to identify the downstream targets of ZNF582-AS1. rRNA MeRIP-seq and MeRIP-qRT-PCR were utilized to examine the N(6)-methyladenosine modification status. Western blot and immunohistochemistry assays were used to determine the protein expression level. Results ZNF582-AS1 was downregulated in ccRCC, and decreased ZNF582-AS1 expression was significantly correlated with advanced tumor stage, higher pathological stage, distant metastasis and poor prognosis. Decreased ZNF582-AS1 expression was caused by DNA methylation at the CpG islands within its promoter. ZNF582-AS1 overexpression inhibited cell proliferative, migratory and invasive ability, and increased cell apoptotic rate in vitro and in vivo. Mechanistically, we found that ZNF582-AS1 overexpression suppressed the N(6)-methyladenosine modification of MT-RNR1 by reducing rRNA adenine N(6)-methyltransferase A8K0B9 protein level, resulting in the decrease of MT-RNR1 expression, followed by the inhibition of MT-CO2 protein expression. Furthermore, MT-RNR1 overexpression reversed the decreased MT-CO2 expression and phenotype inhibition of ccRCC induced by increased ZNF582-AS1 expression. Conclusions This study demonstrates for the first time that ZNF582-AS1 functions as a tumor suppressor gene in ccRCC and ZNF582-AS1 may serve as a potential biomarker and therapeutic target of ccRCC.

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Lactobacillus reuteri BM53-1 Produces a Compound That Inhibits Sticky Glucan Synthesis by Streptococcus mutans

Microorganisms ◽

10.3390/microorganisms9071390 ◽

2021 ◽

Vol 9 (7) ◽

pp. 1390

Author(s):

Masafumi Noda ◽

Naho Sugihara ◽

Yoshimi Sugimoto ◽

Ikue Hayashi ◽

Sachiko Sugimoto ◽

...

Keyword(s):

Antibacterial Activity ◽

Lactic Acid ◽

Dental Caries ◽

Lactobacillus Reuteri ◽

Early Stage ◽

Pcr Analysis ◽

Cariogenic Bacteria ◽

Qrt Pcr ◽

Actinidia Polygama ◽

Glucan Synthesis

Cariogenic bacteria, such as Streptococcus (S.) mutans and S. sobrinus, produce insoluble and sticky glucans as a biofilm material. The present study demonstrates that a lactic acid bacterium (LAB) named BM53-1 produces a substance that inhibits the sticky glucan synthesis. The BM53-1 strain was isolated from a flower of Actinidia polygama and identified as Lactobacillus reuteri. The substance that inhibits sticky glucan synthesis does not exhibit antibacterial activity against S. mutans. The cariogenic S. mutans produces glucans under the control of three glucosyltransferase (GTF) enzymes, named GtfB, GtfC, and GtfD. Although GtfB and GtfC produce insoluble glucans, GtfD forms soluble glucans. Through quantitative reverse-transcriptional (qRT)-PCR analysis, it was revealed that the BM53-1-derived glucan-production inhibitor (GI) enhances the transcriptions of gtfB and gtfC genes 2- to 7-fold at the early stage of cultivation. However, that of gtfD was not enhanced in the presence of the GI, indicating that the glucan stickiness produced by S. mutans was significantly weaker in the presence of the GI. Our result demonstrates that Lb. reuteri BM53-1 is useful to prevent dental caries.

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Comprehensive Analysis of the Expression of Key Genes Related to Hippo Signaling and Their Prognosis Impact in Ovarian Cancer

Diagnostics ◽

10.3390/diagnostics11020344 ◽

2021 ◽

Vol 11 (2) ◽

pp. 344

Author(s):

Paul Kubelac ◽

Cornelia Braicu ◽

Lajos Raduly ◽

Paul Chiroi ◽

Andreea Nutu ◽

...

Keyword(s):

Ovarian Cancer ◽

Cancer Progression ◽

Prognostic Value ◽

Prognostic Significance ◽

Expression Level ◽

Additional Patient ◽

Tumor Control ◽

Hippo Signaling ◽

Patient Cohort ◽

Qrt Pcr

The Hippo signaling pathway, one of the most conserved in humans, controlling dimensions of organs and tumor growth, is frequently deregulated in several human malignancies, including ovarian cancer (OC). The alteration of Hippo signaling has been reported to contribute to ovarian carcinogenesis and progression. However, the prognostic roles of individual Hippo genes in OC patients remain elusive. Herein we investigated the expression level and prognostic value of key Hippo genes in OC using online databases, followed by a qRT-PCR validation step in an additional patient cohort. Using the GEPIA database, we observed an increased level for TP53 and reduced expression level for LATS1, LATS2, MST1, TAZ, and TEF in tumor tissue versus normal adjacent tissue. Moreover, LATS1, LATS2, TP53, TAZ, and TEF expression levels have prognostic significance correlated with progression-free survival. The qRT-PCR validation step was conducted in an OC patient cohort comprising 29 tumor tissues and 20 normal adjacent tissues, endorsing the expression level for LATS1, LATS2, and TP53, as well as for two of the miRNAs targeting the TP53 gene, revealing miR-25-3p upregulation and miR-181c-5p downregulation. These results display that there are critical prognostic value dysregulations of the Hippo genes in OC. Our data demonstrate the major role the conserved Hippo pathway presents in tumor control, underlying potential therapeutic strategies and controlling several steps modulated by miRNAs and their target genes that could limit ovarian cancer progression.

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Identification of miRNAs and Their Response to Cold Stress in Astragalus Membranaceus

Biomolecules ◽

10.3390/biom9050182 ◽

2019 ◽

Vol 9 (5) ◽

pp. 182 ◽

Cited By ~ 6

Author(s):

Merhaba Abla ◽

Huigai Sun ◽

Zhuyun Li ◽

Chunxiang Wei ◽

Fei Gao ◽

...

Keyword(s):

Cold Stress ◽

Redox Homeostasis ◽

Astragalus Membranaceus ◽

Pcr Analysis ◽

Small Rna Sequencing ◽

Factors Affecting ◽

Gene Regulation Network ◽

Qrt Pcr ◽

Yield And Quality ◽

Regulation Network

Astragalus membranaceus is an important medicinal plant widely cultivated in East Asia. MicroRNAs (miRNAs) are endogenous regulatory molecules that play essential roles in plant growth, development, and the response to environmental stresses. Cold is one of the key environmental factors affecting the yield and quality of A. membranaceus, and miRNAs may mediate the gene regulation network under cold stress in A. membranaceus. To identify miRNAs and reveal their functions in cold stress response in A. membranaceus, small RNA sequencing was conducted followed by bioinformatics analysis, and quantitative real time PCR (qRT-PCR) analysis was performed to profile the expression of miRNAs under cold stress. A total of 168 conserved miRNAs belonging to 34 families and 14 putative non-conserved miRNAs were identified. Many miRNA targets were predicted and these targets were involved in diversified regulatory and metabolic pathways. By using qRT-PCR, 27 miRNAs were found to be responsive to cold stress, including 4 cold stress-induced and 17 cold-repressed conserved miRNAs, and 6 cold-induced non-conserved miRNAs. These cold-responsive miRNAs probably mediate the response to cold stress by regulating development, hormone signaling, defense, redox homeostasis, and secondary metabolism in A. membranaceus. These cold-corresponsive miRNAs may be used as the candidate genes in further molecular breeding for improving cold tolerance of A. membranaceus.

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Small Non-Coding RNAome of Ageing Chondrocytes

International Journal of Molecular Sciences ◽

10.3390/ijms21165675 ◽

2020 ◽

Vol 21 (16) ◽

pp. 5675

Author(s):

Panagiotis Balaskas ◽

Jonathan A. Green ◽

Tariq M. Haqqi ◽

Philip Dyer ◽

Yalda A. Kharaz ◽

...

Keyword(s):

Quantitative Polymerase Chain Reaction ◽

Differential Expression Analysis ◽

Cartilage Tissue ◽

Pcr Analysis ◽

Small Rna Sequencing ◽

High Grade ◽

Human Cartilage ◽

Fragment Analysis ◽

Qrt Pcr ◽

Age Related

Ageing is a leading risk factor predisposing cartilage to osteoarthritis. However, little research has been conducted on the effect of ageing on the expression of small non-coding RNAs (sncRNAs). RNA from young and old chondrocytes from macroscopically normal equine metacarpophalangeal joints was extracted and subjected to small RNA sequencing (RNA-seq). Differential expression analysis was performed in R using package DESeq2. For transfer RNA (tRNA) fragment analysis, tRNA reads were aligned to horse tRNA sequences using Bowtie2 version 2.2.5. Selected microRNA (miRNAs or miRs) and small nucleolar RNA (snoRNA) findings were validated using real-time quantitative Polymerase Chain Reaction (qRT-PCR) in an extended cohort of equine chondrocytes. tRNA fragments were further investigated in low- and high-grade OA human cartilage tissue. In total, 83 sncRNAs were differentially expressed between young and old equine chondrocytes, including miRNAs, snoRNAs, small nuclear RNAs (snRNAs), and tRNAs. qRT-PCR analysis confirmed findings. tRNA fragment analysis revealed that tRNA halves (tiRNAs), tiRNA-5035-GluCTC and tiRNA-5031-GluCTC-1 were reduced in both high grade OA human cartilage and old equine chondrocytes. For the first time, we have measured the effect of ageing on the expression of sncRNAs in equine chondrocytes. Changes were detected in a number of different sncRNA species. This study supports a role for sncRNAs in ageing cartilage and their potential involvement in age-related cartilage diseases.

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