Sagittaria sagittifolia L is an important bulb vegetable that has high nutritional and medical value. Bulb formation and development are crucial to Sagittaria sagittifolia; however, its sucrose metabolism is poorly understood and there are a lack of sufficient transcriptomic and genomic data available to fully understand the molecular mechanisms underlying bulb formation and development as well as the bulb transcriptome. Five cDNA libraries were constructed at different developmental stages and sequenced using high-throughput Illumina RNA sequencing. From approximately 63.53 Gb clean reads, a total of 60,884 unigenes, with an average length of 897.34 bp and N50 of 1.368 kb, were obtained. A total of 36,590 unigenes were successfully annotated using five public databases. Across different developmental stages, 4195, 827, 832, 851, and 1494 were differentially expressed in T02, T03, T04, T05, and T06 libraries, respectively. Gene ontology (GO) analysis revealed several differentially-expressed genes (DEGs) associated with catalytic activity, binding, and transporter activity. The Kyoto encyclopedia of genes and genomes (KEGG) revealed that these DEGs are involved in physiological and biochemical processes. RT-qPCR was used to profile the expression of these unigenes and revealed that the expression patterns of the DEGs were consistent with the transcriptome data. In this study, we conducted a comparative gene expression analysis at the transcriptional level using RNA-seq across the different developmental stages of Sagittaria sagittifolia. We identified a set of genes that might contribute to starch and sucrose metabolism, and the genetic mechanisms related to bulblet development were also explored. This study provides important data for future studies of the genetic and molecular mechanisms underlying bulb formation and development in Sagittaria sagittifolia.
Interferon-gamma-inducible protein 30 (IFI30) is an IFN-gamma-inducible protein that is involved in MHC class II-restricted antigen processing and MHC class I-restricted cross-presentation pathways of adaptive immunity. The present study aimed to investigate the effects of porcine IFI30 expression on PRRSV proliferation in host cells. MARC-145 cells and pig Sertoli (ST) cells were infected with PRRSV after transfection with porcine IFI30 expression vectors and an empty vector. PRRSV copy numbers were analyzed by absolute real-time quantitative PCR, and the results showed that porcine IFI30 expression could significantly inhibit PRRSV transcription. Western blot analysis also determined that IFI30 expression could reduce the production of PRRSV M protein. Flow cytometric analysis indicated that the apoptosis of MARC-145 cells, which are non-porcine but highly permissive to PRRSV cells, was significantly decreased in the IFI30 expression group. In porcine ST cells, apoptosis was significantly increased in IFI30 knockdown cells but not in IFI30-overexpressing cells (**p < 0.01). In conclusion, porcine IFI30 expression may inhibit PRRSV proliferation and host cell apoptosis in vitro.
We looked for differentially expressed MicroRNAs (miRNAs) in Immunoglobulin A nephropathy (IgAN). Forty-eight miRNAs were identified through the initial screening phase (2 IgAN pools vs. 1 normal control (NC) pool) using quantitative reverse transcription polymerase chain reaction (qRT-PCR) based Exiqon panel (miRCURY-Ready-to-Use-PCR-Human-panel-I + II-V-1.M). By qRT-PCR, these miRNAs were further assessed in the training (32 IgAN VS. 31 NCs) and testing stages (51 IgAN VS. 51 NCs). The renal pathological lesions of patients with IgAN were evaluated according to Lees grading system. We discovered a plasma miRNA signature including four up-regulated miRNAs (miR-148a-3p, miR-150-5p, miR-20a-5p and miR-425-3p) and the areas under the receiver operating characteristic (ROC) curve (AUC) were 0.80 and 0.76 for the training and testing stage, respectively. The expression of the four miRNAs in IgAN grade I-II subgroups (according to Lees grading system) was obviously higher than that in IgAN grade III-V (P < .05). In summary, the plasma expression of miR-148a-3p, miR-150-5p, miR-20a-5p and miR-425-3p were up-regulated in patients with IgAN, especially the early-stage disease. Further studies are needed to explore the roles of the four miRNAs in the pathogenesis and progression of IgAN.
Anther cuticle and pollen exine are two elaborated lipid-soluble barriers protecting pollen grains from environmental and biological stresses. However, less is known about the mechanisms underlying the synthesis of these lipidic polymers. Here, we identified a no-pollen male-sterility mutant cyp703a3-3 from the indica restorer line Zhonghui 8015 (Zh8015) mutant library treated with 6 Coy-ray radiation. Histological analysis indicated that cyp703a3-3 underwent abnormal tapetal cells development, produced few orbicules and secreted less sporopollenin precursors to anther locule, as well as cutin monomers on anther. Genetic analysis revealed that cyp703a3-3 was controlled by a single recessive gene. Map-based cloning was performed to narrow down the mutant gene to a 47.78-kb interval on the chromosome 8 between two markers S15-29 and 515-30. Sequence analysis detected three bases (GAA) deletion in the first exon of LOC_Os08g03682, annotated as CYP703A3 with homologous sequences related to male sterility in Arabidopsis, causing the Asparagine deletion in the mutant site. Moreover, we transformed genomic fragment of CYP703A3 into cyp703a3-3, which male-sterility phenotype was recovered. Both the wild-type and cyp703a3-3 mutant 3D structure of CYP703A3 protein were modeled. Results of qPCR suggested CYP703A3 mainly expressed in anthers with greatest abundance at microspore stage, and genes involved in sporopollenin precursors formation and transportation, such as GAMYB, TDR, CYP70482, DPW2, OsABCG26 and OsABCG15, were significantly reduced in cyp703a3-3. Collectively, our results further elaborated CYP703A3 plays vital role in anther cuticle and pollen exine development in rice (Oryza sativa L.).
Long noncoding RNAs is a novel class of RNA molecules, which is closely related to the occurrence and development of human disease. Recent studies have highlighted the importance of MEG3 in angiogenesis and the maintenance of normal function of vascular endothelial cells. However, whether MEG3 contributes to human endothelial cell angiogenesis as well as potential mechanisms are largely unknown. In this work, we found that the high expression level of MEG3 in primary HUVEC was controlled by DNA methylation, and its expression in HUVEC was regulated by HIF-1 alpha under hypoxia condition. Meanwhile, we discovered that knockdown of MEG3 significantly suppressed VEGFR2 mRNA level, but had no influence on gene expression of VEGFR1, Notch1, DLL4 and Hes1. MEG3 reduction also suppressed VEGF-induced endothelial migration and angiogenesis. Furthermore, MEG3 knockdown reduced the tube formation and the spheroid sprouting of primary HUVEC under normoxic and hypoxic conditions. Altogether, MEG3 regulated by HIF-1 alpha is required to maintain VEGFR2 expression in endothelial cells and plays a vital role for VEGFA-mediated endothelial angiogenesis.
The doublesex and its homologue genes are important regulators of sexual differentiation which are conserved among animal kingdom. In the present study, we reported a doublesex gene (designated as FcDsx) identified from the Chinese shrimp F. chinensis. The gene structure, nucleotide and deduced amino acid sequences of FcDsx were characterized. The results showed that the deduced amino acid sequence of FcDsx had the common features of Dsx proteins, including a doublesex/male abnormal 3 (DM) domain, an oligomerization domain and a predicted monopartite nuclear localization signal. The expression patterns of FcDsx in different tissues and developmental stages were detected. FcDsx exhibited a sex-biased expression patterns in different tissues and its expression level increased along with developmental stages. In addition, its regulation on the expression of FcIAG, a gene important for sexual differentiation of male crustacean, was also analyzed. Putative Dsx binding site was identified on the promoter region of FcIAG and knockdown of FcDsx could reduce the expression of FcIAG, which suggested that FcDsx might be the upstream regulator of FcIAG. The present data indicated that FcDsx gene might involve in shrimp sexual differentiation process.
Objective: The present study aimed to investigate the molecular mechanism underlying congenital obstructive nephropathy (CON). Methods: The microarray dataset GSE70879 was downloaded from the Gene Expression Omnibus, including 3 kidney samples of megabladder mice and 4 control kidneys. Using this dataset, differentially expressed miRNAs (DEMs) were identified between the kidney samples from megabladder mice and controls, followed by identification of the target genes for these DEMs and construction of a DEM and target gene interaction network. Additionally, the target genes were subjected to Gene Ontology and pathway enrichment analyses, and were used for construction of a protein-protein interaction (PPI) network. Finally, regulatory networks were constructed to analyze transcription factors for the key miRNAs. Results: From 17 DEMs identified between kidney samples of megabladder mice and controls, 3 key miRNAs were screened, including mmu-miR-150-5p, mmu-miR-374b-5p and mmu-miR-126a-5p. The regulatory networks identified vascular endothelial growth factor A (Vegfa) as the common target gene of mmu-miR-150-5p and five transcription factors, including nuclear receptor subfamily 4, group A, member 2 (Nr4a2), Jun dimerisation protein 2 (Jdp2), Kruppel-like factor 6 (Klf6), Neurexophilin-3 (Nxph3) and RNA binding motif protein 17 (Rbm17). The gene encoding phosphatase and tensin homolog (Pten) was found to be co-regulated by mmu-miR-374b-5p and high mobility group protein A1 (Hmga1), whereas the kirsten rat sarcoma viral oncogene (Kras) was identified as a common target gene of mmu-miR-126a-5p and paired box 6 (Pax6). Conclusions: In summary, the above-listed key miRNAs, transcription factors and key genes may be involved in the development of CON.
Growing evidence shows that long non-coding RNAs (IncRNAs) have been wildly verified to modulate multiple tumorigenesis, especially lung adenocarcinoma. In present study, we aim to investigate the role of IncRNA LINC00319 in the lung adenocarcinoma carcinogenesis. We observed that increased expression of LINC00319 in lung adenocarcinoma tissues and cells in comparison to their corresponding controls. Moreover, the aberrant overexpression of LINC00319 indicated the poor prognosis of lung adenocarcinoma patients. Silence of LINC00319 was able to repress lung adenocarcinoma cell growth in vitro. Rescue assay was performed to further confirm that LINC00319 contributed to lung adenocarcinoma progression by regulating miR-450b-5p/EZH2 signal pathway. Taken together, our study discovered the oncogenic role of LINC00319 in clinical specimens and cellular experiments, showing the potential LINC00319/miR-450b-5p/EZH2 pathway. This results and findings provide a novel insight for lung adenocarcinoma tumorigenesis.
The domestic yak (Bos grunniens) is a culturally important animal that lives at high altitude and is farmed by Tibetan herders for its meat, milk, and other animal by-products. Within the animal, adipose tissue is an important store and source of energy and is used to maintain adequate body temperature during the extended cold seasons. Exploring the biomolecular role of microRNAs (miRNAs) in the regulation of growth, development, and metabolism of yak adipocytes may provide valuable insights into the physiology of adipogenesis in the yak. This study investigated whether and how miR-200a (a miRNA recently reported to promote adipogenesis in ST2 bone marrow stromal cells) regulates adipocyte differentiation in the yak. Expression levels of miR-200a gradually increased during day 0 to day 8 of adipocyte differentiation, and transfection of adipocytes with miR-200a enhanced lipid accumulation and triglyceride content compared to control (un-transfected) adipocytes. We additionally verified (using qRT-PCR analysis) that miR-200a increased the expression of adipocyte-specific genes involved in lipogenic transcription (PPAR gamma, ELVOL, and C/EBP alpha), fatty acid synthesis (ACC, ACS, SCD, and FAS), and fatty acid transport (DGAT, LPL, and FABP4). We also found that transfection of adipocytes with miR200a resulted in suppression of the levels of noncanonical Wnt signaling transcription factors (Wnt5a, TAKI, and NLK). These results indicate that miRNA-200a plays an important role in promoting yak adipocyte differentiation that may operate via the suppression of noncanonical Wnt signaling.
Lysine specific demethylase 513 gene (KDM5B, also known as JARIDIB or PLU-1), encoding an enzyme of the lysine-specific histone demethylase family, has been reported to regulate androgen receptor transcriptional activity and male reproduction. To fully study the expression characteristics and genetic effects of pig KDM5B gene, the objective of this study was to investigate the mRNA expression profiles of KDM5B among different tissues and testicular cells (spermatogonia stem cells, SSCs; sertoli cells, SCs; leydig cells, LCs), as well as to explore the insertion/deletion (indel) variations of this gene. Expression analysis results revealed that the KDM5B gene was highly expressed in testis than other tissues in 7-day-old piglet (P < 0.01). In particular, this gene was highest expressed in testis of adult boar (P < 0.01). Furthermore, the KDM5B mRNA expression level in SSCs was significantly higher than those in LCs and SCs (P < 0.01). Besides, a 35-bp indel was first verified in the intron 11 of pig KDM5B gene, and the animals with insertion/insertion genotype exhibited superior testicular weight and testicular short perimeter than those with other genotypes (P < 0.05) in 40-day-old Landrace pigs. Together, these findings suggest that KDM5B plays a crucial role in male fertility and the 35-bp indel can be used for marker-assisted selection of boar.
Increasing amounts of evidence have indicated that non-coding RNAs (ncRNAs) have important regulatory potential in various biological processes. However, the contribution of ncRNAs, especially long non-coding RNAs (IncRNAs) to drug induced steatosis remain largely unknown. The aim of this study is to investigate miRNA, IncRNA and mRNA expression profiles and their potential roles in the process of drug induced steatosis. Microarray expression profiles of miRNAs, IncRNAs and mRNAs were determined in dexamethasone treated HepG2 cell as well as control cell. Differential expression, pathway and gene network analyses were developed to identify possible functional RNA molecules in dexamethasone induced steatosis. Compared with control HepG2 cell, 652 IncRNAs (528 up-regulated and 124 down-regulated), 655 mRNAs (527 upregulated and 128 down regulated) and 114 miRNAs (55 miRNAs up-regulated and 59 down-regulated) were differentially expressed in dexamethasone treated HepG2 cell. Pathway analysis showed that the fatty acid biosynthesis, insulin resistance, PPAR signaling pathway, regulation of lipolysis in adipocytes, carbohydrate digestion and absorption, steroid hormone biosynthesis signaling pathways had a close relationship with dexamethasone induced steatosis. 10 highly dysregulated mRNAs and 20 miRNAs, which are closely related to lipid metabolism, were identified and validated by PCR, which followed by ceRNA analysis. CeRNA network analysis identified 5 lipid metabolism related genes, including CYP7A1, CYP11A1, PDK4, ABHD5, ACSL1. It also identified 12 miRNAs (miR-23a-3p, miR-519d-3p, miR-4328, miR-15b-5p etc.) and 177 IncRNAs (ENST00000508884, ENST00000608794, ENST00000568457 etc.). Our results provide a foundation and an expansive view of the roles and mechanisms of ncRNAs in dexamethasone induced steatosis.
Osteosarcoma (OS) is a common primary malignancy in children and adolescents with relative high survival rate after chemotherapy. While, the toxicity of chemotherapy and personalized different response to chemotherapy makes it difficult for the selection of therapeutics and improvement of diagnosis. In this study, we conducted a combined analysis of two types of microarray datasets (gene expression and DNA methylation) from the Gene Expression Omnibus (GEO). Differential methylation sites (DMS) were identified by the IMA package and differential expression genes (DEGs) were screened out via the limma package. A total of 11,242 DMS (corresponding to 3080 genes (DMGs)) and 337 DEGs, with 40 overlaps (OS genes) between DEGs and DMGs, were identified. Enriched functions of OS genes were obtained through the Database for Annotation, Visualization and Integrated Discovery (DAVID). The OS genes were mainly enriched in the biological processes related to inflammatory/immune response and Pertussis pathways and Hematopoietic cell lineage pathways. Besides, OS specific disease network was obtained, and found that UBS and NRF1 were regulated by multiple OS genes. Kaplan Meier analysis of OS genes identified BHMT2, DOCK2, DNALI1 and RIPK3 as significant OS survival related genes. SEMA3A and PRAME are included in the 40 OS genes and within the top 10 most up-regulated DEGs. Their expression changes were further validated in U2OS osteosarcoma cell lines and hOB normal cell lines through quantitative PCR (qPCR) and consistent result with microarray analysis was obtained. Based on this study, some novel targets were identified for OS, which would be helpful in its early diagnosis and treatment.
Objective: High altitude pulmonary edema (HAPE) is a life threatening condition occurring in otherwise healthy individuals who rapidly ascend to high altitude. However, the molecular mechanisms of its pathophysiology are not well understood. The objective of this study is to evaluate differential gene expression in patients with HAPE during acute illness and subsequent recovery. Methods: Twenty-one individuals who ascended to an altitude of 3780 m were studied, including 12 patients who developed HAPE and 9 matched controls without HAPE. Whole-blood samples were collected during acute illness and subsequent recovery for analysis of the expression of hypoxia-related genes, and physiologic and laboratory parameters, including mean pulmonary arterial pressure (mPAP), heart rate, blood pressure, and arterial oxygen saturation (SpO(2)), were also measured. Results: Compared with control subjects, numerous hypoxia-related genes were up-regulated in patients with acute HAPE. Gene network analyses suggested that HIF-1 alpha played a central role in acute HAPE by affecting a variety of hypoxia-related genes, including BNIP3L, VEGFA, ANGPTL4 and EGLN1. Transcriptomic profiling revealed the expression of most HAPE-induced genes was restored to a normal level during the recovery phase except some key hypoxia response factors, such asBNIP3L, EGRJ, MMP9 and VEGF, which remained persistently elevated. Conclusions: Differential expression analysis of hypoxia-related genes revealed distinct molecular signatures of HAPE during acute and recovery phases. This study may help us to better understand HAPE pathogenesis and putative targets for further investigation and therapeutic intervention.
This study aimed to explore crucial genes, transcription factors (TFs), and microRNAs (miRNAs) associated with the effects of curcumin against hepatocellular carcinoma (HCC). We downloaded data (GSE59713) from Gene Expression Omnibus to analyze differentially expressed genes (DEGs) between curcumin-treated and untreated HCC cell lines. Then, we identified the disease ontology (DO) and functional enrichment analysis of these DEGs and analyzed their protein-protein interactions (PPIs). Additionally, we constructed TF-target gene and miRNA-target gene regulatory networks and explored the potential functions of these DEGs. Finally, we detected the expression of CDKN1A, CTGF, LEF1 TF and MIR-19A regulated by curcumin in PLC/PRF/5 cells using RT-PCR. In total, 345 upregulated and 212 downregulated genes were identified. The main enriched pathway of upregulated genes was the TNF signaling pathway. The downregulated genes were significantly enriched in TGF-beta signaling pathway. In addition, most DEGs were significantly enriched in DO terms such as liver cirrhosis, hepatitis, hepatitis C and cholestasis (eg., CTGF). In the constructed PPI network, CDKN1A and CTGF were the key proteins. Moreover, LEF1, CDKN1A, and miR-19A that regulated CTGF were highlighted in the regulatory networks. Furthermore, the expression of CDKN1A, CTGF, LEF1 TF and miR-19A regulated by curcumin in PLC/PRF/5 cells was consistent with the aforementioned bioinformatics analysis results. To conclude, curcumin might exert its protective effects against HCC tumorigenesis by downregulating LEF1 and downregulating CTGF regulated by MIR-19A and upregulating CDKN1A expression.
Background: Genome-wide association study (GWAS) studies have showed that single nucleotide polymorphisms (SNPs) in OCA2 gene were associated with the survival of breast cancer patients treated with adjuvant chemotherapy. To further explain the association between OCA2 SNPs and breast cancer survival, we investigated the predictive value of rs4778137 located in OCA2 in local advanced breast cancer patients receiving neoadjuvant chemotherapy. Patients and methods: A case-cohort with 150 breast cancer patients was performed to evaluate the effects of the OCA2 rs4778137 on breast cancer survival. The association between rs4778137 genotypes and pathological complete response (pCR, defined that the postoperative pathology indicating no residual invasive breast cancer in the breast or the axillary lymph node) were analyzed. Logistic regression analysis was performed to identify the independent predictors of pCR. Survival was assessed by Kaplan-Meier method and Cox regression analysis according to the rs4778137 genotypes. Results: The differences between pCR and the rs4778137 genotypes were statistically significant (p < 0.05). The patients with genotype GG harbored a better disease-free survival (HR: 2.358, p = 0.000) and overall survival (HR: 1.578, p = 0.008) than the patients with genotype CC in rs4778137. The further Univariate and Multivariate survival analysis revealed that SNP rs4778137 was an independent predictive factor of disease-free survival (p = 0.000/p = 0.001) and overall survival (p = 0.006/p = 0.045). Conclusion: The OCA2 rs4778137 may be a predictor for the clinical response and survival in local advanced breast cancer patients who received neoadjuvant chemotherapy.