Macromolecule Biosynthetic Process Research Articles

GAPVD1 Promotes the Proliferation of Triple-Negative Breast Cancer Cells by Regulating the ERK/MAPK Signaling Pathway.

Triple-Negative Breast Cancer (TNBC) accounts for 15-20% of all breast cancers and approximately 50% of breast cancer deaths. Chemotherapy remains the mainstay of systemic treatment due to the lack of effective therapy targets. Thus, more studies are urgently needed to identify new therapeutic targets in TNBC patients. GAPVD1 expression and prognosis value in breast cancer samples were explored in The Cancer Genome Atlas database (TCGA). GAPVD1 knockdown and overexpression TNBC cell lines were constructed. CCK-8 and colony formation assays were performed to detect cell viability. Flow cytometry analysis was performed to detect cell cycle variation. Western blotting was conducted to determine the levels of target genes. Finally, an enrichment analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed. GAPVD1 is overexpressed in breast cancer tissues and predicts poor prognosis. In vitro experiments demonstrated that GAPVD1 is correlated with cell proliferation and the cell cycle of TNBC cells. Mechanistically, alteration in GAPVD1 expression was found to be associated with cell cycle-related proteins PCNA, Cyclin A, and the activity of the ERK/MAPK signaling pathway. Consistent with these findings, enrichment analysis of GAPVD1-involving partners and signaling pathways revealed that the cellular biosynthetic process, macromolecule biosynthetic process, and cell cycle signaling are related to GAPVD1. In vivo experiment demonstrated that GAPVD1 inhibition impedes tumor growth and expression of cell cyclerelated proteins. Taken together, our results indicate that GAPVD1 may participate in TNBC cell growth by regulating the cell cycle and ERK/MAPK signaling pathway.

O-306 Sperm DNA methylome changes in testicular cancer patients following chemotherapy treatment

Abstract Study question Does chemotherapy (CT) treatment for testicular cancer induce sperm methylome changes? Summary answer The analysis of 27 pre- and post-CT samples from testicular cancer survivors identified 65 differentially methylated regions. What is known already Research has shown that cancer treatment can cause DNA damage and aneuploidy in sperm, even years after the treatment. However, there is limited information available on the impact of CT on the sperm epigenome, and most of the information has been obtained from rodent models. Two recent studies, one involving one patient and the other involving three, have investigated this topic in humans. Both studies have suggested that cancer survivors may experience changes in sperm’s DNA methylation following oncological treatment. Study design, size, duration This is a multicenter prospective study. We included 27 patients [31.23 years (CI 95% 29.29-33.17)] diagnosed with testicular cancer who cryopreserved semen before CT (2009-2020). After receiving oncological treatment, these patients recovered their spermatogenesis. Patients who agreed to participate in this IRB-approved study, donated a second semen sample after their recovery, which was also frozen. Participants/materials, setting, methods Semen samples were thawed; sperm DNA was isolated and subsequently hybridized using the Human Infinitum DNA Methylation 850K EPIC BeadChip array platform (Illumina). A paired differential methylation analysis was performed between pre-CT and post-CT samples from the same patient, fitting a linear model of the methylated positions and regions. Minfi and DMRcate R packages were used to find differentially methylated positions (DMPs) and regions (DMRs). Subsequently, the functional impact of involved genes was evaluated. Main results and the role of chance DNA methylation data were available for the 54 sperm samples (27 collected before CT and 27 after CT treatment). Results were controlled by age and time elapsed between the cryopreservation of pre-CT and post-CT samples, which was 4.01 years (CI 95%: 3.42-5.64). After filtering for initial QC, the assessment was done on 445.000 CpGs per sample. The analysis revealed that a total of 228 CpG positions showed a tendency towards statistical significance. DMRcate identified 65 DMRs (40 hypomethylated and 25 hypermethylated) that reached a smoothed FDR p-value < 0.05 when compared post-CT samples to pre-CT within the same patient. We observed that 61 genes were located within DMRs, being TBC1D20, EN1, HOXB1 and SCNN1A the ones within the most significant DMRs (min-smoothed-FDR < 0.001). The functional enrichment analysis of putatively affected genes revealed that the biological processes more affected were: Regulation of macromolecule biosynthetic process, hormone receptor binding, signaling pathways regulating pluripotency of stem cells and pathways in cancer. Limitations, reasons for caution Epigenetics may be affected during cryopreservation and thawing. To ensure comparability, we also froze the samples after undergoing the CT process. Due to the high-throughput nature of the study and the limited access to samples, we only achieved marginal significance. Therefore, our results should be further validated. Wider implications of the findings This is the largest study suggesting distinct sperm DNA methylation signatures in testicular cancer survivors. Our findings emphasize the importance of counseling cancer patients in pretreatment sample collection and may explain the reported increase in birth defects after treatment. Trial registration number NOT APPLICABLE

4D label-free proteomics analysis of oxygen-induced retinopathy with or without anti-VEGF treatment

Oxygen-induced retinopathy (OIR) animal model is widely used for retinopathy of prematurity (ROP) researches. The purpose of this study was to identify proteins and related pathways of OIR with or without anti-vascular endothelial growth factor (VEGF) treatment, for use as biomarkers in diagnosing and treating ROP. Nine samples were subjected to proteomic analysis. Retina specimens were collected from 3 OIR mice, 3 OIR mice with anti-VEGF treatment and 3 normal mice (control group). Liquid chromatography-tandem mass spectrometry analysis was performed using the 4D label-free technique. Statistically significant differentially expressed proteins, gene ontology (GO) terms, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway representations, InterPro (IPR) and protein interactions were analyzed. In total, 4585 unique proteins were identified as differentially expressed proteins (DEPs). Enrichment analysis of the GO and KEGG indicated functional clusters related to peptide biosynthetic and metabolic process, cellular macromolecule biosynthetic process and nucleic acid binding in OIR group. For anti-VEGF treatment group, DEPs were clustered in DNA replication, PI3K/Akt signaling pathway and Jak/STAT signaling pathway. Proteomic profiling is useful for the exploration of molecular mechanisms of OIR and mechanisms of anti-VEGF treatment. These findings may be useful for identification of novel biomarkers for ROP pathogenesis and treatment.

Transcriptomic Analysis Reveals That Retinal Neuromodulation Is a Relevant Mechanism in the Neuroprotective Effect of Sitagliptin in an Experimental Model of Diabetic Retinopathy

Synaptic dysfunction and neuronal damage have been extensively associated with diabetic retinopathy (DR). Our group evidenced that chronic hyperglycemia reduces the retinal expression of presynaptic proteins, which are crucial for proper synaptic function. The aim of the study was to explore the effect of topically administered sitagliptin, an inhibitor of the enzyme dipeptidyl peptidase-4, on the retinal expression patterns of an experimental model of DR. Transcriptome analysis was performed, comparing the retinas of 10 diabetic (db/db) mice randomly treated with sitagliptin eye drops (10 mg/mL) twice daily and the retinas of 10 additional db/db mice that received vehicle eye drops. Ten non-diabetic mice (db/+) were used as a control group. The Gene Ontology (GO) and Reactome databases were used to perform the gene set enrichment analysis (GSEA) in order to explore the most enriched biological pathways among the groups. The most differentiated genes of these pathways were validated through quantitative RT-PCR. Transcriptome analysis revealed that sitagliptin eye drops have a significant effect on retinal expression patterns and that neurotransmission is the most enriched biological process. Our study evidenced enriched pathways that contain genes involved in membrane trafficking, transmission across chemical synapses, vesicle-mediated transport, neurotransmitter receptors and postsynaptic signal transmission with negative regulation of signaling as a consequence of neuroprotector treatment with sitagliptin. This improves the modulation of the macromolecule biosynthetic process with positive regulation of cell communication, which provides beneficial effects for the neuronal metabolism. This study suggests that topical administration of sitagliptin ameliorates the abnormalities on presynaptic and postsynaptic signal transmission during experimental DR and that this improvement is one of the main mechanisms behind the previously demonstrated beneficial effects.

Mass Spectrometry-Based Quantitative Proteomics Analysis for Better Understanding of Telomeric Zinc Finger-Associated Protein-Induced Pathogenesis in Cardiomyocytes.

Telomere length is highly related to cardiovascular diseases. Telomeric zinc finger-associated protein (TZAP) directly binds to telomeric TTAGGG repeats via zinc finger domains and triggers the initiation of the telomere trimming process. However, proteomics analysis of TZAP in cardiomyocytes is slightly unknown. In our study, TZAP was overexpressed by adenovirus transfection in cultured H9c2 cardiomyocytes, and then mass spectrometry-based quantitative proteomics research strategies, including Gene Ontology analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, subcellular localizations, predicted functional domains, and protein-protein interaction (PPI) analysis, were performed to explore TZAP-induced potential pathogenesis in cardiomyocytes. A total of 184 upregulated and 228 downregulated differentially expressed proteins (DEPs) were identified among identified 5693 quantifiable proteins in TZAP-overexpressed cardiomyocytes. These DEPs were mainly distributed in the nucleus, cytoplasm, and plasma membrane. DEPs were enriched in biological processes including cardiac muscle cell contraction, acute inflammatory response, cell-cell junction assembly, and macromolecule biosynthetic process. They were enriched in 9 KEGG pathways, including Hippo signaling pathway, protein digestion and absorption, and cytokine receptor interaction, and enriched in 17 protein domains, including translation initiation factor 1A/IF-1, class I histocompatibility antigen, and zinc finger. PPI analysis indicated that TZAP interacted with NDUFC2, Gja1, and HDAC2. Further, as proteins closely related to cardiovascular function, the mRNA levels of BRD4, Gja1, HDAC2, MAP2K3, Plakophilin 4, and Syndecan 1 significantly decreased, while Trpm7, clusterin, and NDUFC2 remarkably increased in TZAP-overexpressed cardiomyocytes by RT-PCR assay, which were consistent with the proteomics analysis. Collectively, we provided candidate proteins and enrichment pathways in TZAP-overexpressed cardiomyocytes, which need further investigation.

Comparative Transcriptome Analysis Revealed the Freezing Tolerance Signaling Events in Winter Rapeseed (Brassica rapa L.).

Winter rapeseed (Brassica rapa L.) is an important oilseed crop in northwest China. Freezing stress severely limits its production and geographical distribution, and frequent extreme freezing events caused by climate change are increasing the chances of winter freeze-injury. However, the underlying mechanism of B. rapa response to freezing stress remains elusive. Here, B. rapa genome (v3.0) was used as a reference for the comparative transcriptomic analysis of Longyou 6 and Tianyou 2 (strong and weak cold tolerance, respectively) under different freezing stress. Before and after freezing stress, 5,982 and 11,630 unique differentially expressed genes (DEGs) between two cultivars were identified, respectively. After freezing stress, the GO terms in Tianyou 2 were mainly involved in “macromolecule biosynthetic process”, and those in Longyou 6 were involved in “response to stimulus” and “oxidoreductase activity”. Morphological and physiological results indicated that Longyou 6 retained a higher basal freezing resistance than Tinayou 2, and that cold acclimation could strengthen the basal freezing resistance. Freezing stress could activate the MAPK signal cascades, and the phosphorylation level of Longyou 6 showed a higher increase in response to freezing treatment than Tianyou 2. Based on our findings, it was speculated that the cell membrane of B. rapa perceives external signals under freezing stress, which are then transmitted to the nucleus through the cold-activated MAPK cascades and Ca2+-related protein kinase pathway, thus leading to activation of downstream target genes to enhance the freezing resistance of B. rapa.

Differential gene expression analysis for the identification of potential biomarkers for Niemann-Pick type C disease.

Niemann-Pick disease (NP) is a rare neurovisceral lysosomal lipid storage disorder. NPC (NP type C) is characterized by the accumulation of unesterified cholesterol and other lipids within the cell, the function of lysosomes usually is impaired as well as the autophagy flux. To better understand the molecular pathways affected in this disease, we perform a bioinformatic study. We used in silico methods to identify genes affected in this disease. The first approach was an analysis of microarray data from the public archive GEO2, accession number GSE124283. We studied gene expression of skin fibroblast from 55 individuals, 22 healthy controls (13 males and 9 females) and other 23 with NPC (12 males and 11 females), 16 patients suffering from infantile NPC, 5 juvenile NPC and 2 adults. The differential expression analysis was performed using limma package, genes with significative adjusted p value and fold change > 0.5 and < -0.5 were selected, we used topGO package to obtain the Gene Ontology. The analysis revealed that cellular components enriched were vesicle membrane, vacuolar lumen lysosome, cytoplasm vesicle, cytoskeletal fiber, nucleus and nucleoplasm. For molecular function were affected catalytic activity, cadherin binding and enzyme binding, and the biological process enriched were the phagosome maturation, lysosome organization, vesicular transport, apoptotic process, gene expression, proteosome-mediated ubiquitin, cell cycle, cellular macromolecule biosynthetic process and regulation of cellular response to stress. Our results were consistent with changes in cell homeostasis reported from NPC patients. Abnormal protein degradation and metabolism might contribute to the higher level of mortality in this disease. We propose that some of the DEGs that are more related to NPC disease according to our study, could have value as potential biomarkers of this disease helping to unravel the molecular mechanisms involve in this disease.

Insight to new genes with sex‐biased to bony‐tongued fishes: Differentially expressed genes in adult individuals of Arapaima gigas revealed by RNA‐Seq

The Arapaima gigas is an endemic fish to the Amazon basin and with great potential for aquaculture, although its reproduction in captivity is limited by the absence of sexual dimorphism. The demand increase for pirarucu in the Northern region of Brazil, has led to new studies on the reproduction of the species. Thus, we aimed to investigate patterns of sex-biased genes in brain and gonad tissues of A. gigas. A total of of 451,490,808 short sequencing reads were produced using the SOLiD v4 plus sequencing platform. An average of ~30% of sequencing reads were mapped to Asian arowana reference cDNAs, species that belong to the same group of bony-tongue fishes. Gene expression analysis identified 305 sex-biased genes for the brain, with 297 genes up-regulated in females and only eight genes up-regulated in males. The analysis of the gonads identified 120 sex-biased genes, with 110 genes up-regulated in females and only ten genes up-regulated in males. Gene ontology analysis found more than 50 terms enriched for the sex-biased genes in a wide range of pathways suggesting that multiple functions and processes differ between the sexes. Thus, the following areas of interest were approached: sex steroids/hormones, organism development process and macromolecule biosynthetic process, which exhibited sex-biased gene expression; these areas are known to influence sexual development in other species. Lastly, the information obtained in this study concerning the sex-biased genes in the A. gigas species is a contribution to further studies approaching the processes of gonadal development, and downstream molecular mechanisms in species.

Optimizing the program of sequential chemotherapy for advanced gastric cancer based on gene-level CNVs called from ctDNA.

e16068 Background: Tumor evolutionary trade-offs under treatment pressure can be utilized to optimize therapy. We aimed to profile the dynamic pattern of gene-level copy number variations (CNVs) for advance gastric cancer (GCA) patients who underwent sequential chemotherapy and optimized the therapy program. Methods: 27 chemo-naive advanced GCA patients with distant metastasis were enrolled. Peripheral blood samples were collected at different time points, including pre-therapy and disease progression on a specific regimen. Gene-level CNVs were called from ctDNA of blood samples and drug modified score (DMS) was defined as the ratio of CNV value between samples of pre- and post-treatment for each gene. Positive and negative DMS of a gene represented its resistance and sensitivity to a selected regimen, respectively. Then, the best initial regimen was selected and the optimal program of sequential chemotherapy was designed for each patient. Results: DMS of gene was calculated for four chemotherapy regimens, including platinum-based doublet regimen (Pt-2d), oral-fluorouracil prodrug (O-FP), paclitaxel-based regimen (PTX) and irinotecan-based regimen (CPT-11). Based on DMS distribution, resistant and sensitive genes were determined for each regimen through a heuristic method. Functional enrichment analysis showed that resistant genes mainly involved in cellular response to oxygen-containing compound and regulation of cell proliferation, while sensitive genes mainly involved in positive regulation of macromolecule biosynthetic process and regulation of apoptotic process. Then, drug resistant score (DRS, defined as DMS weighted sum of CNV values of resistant genes) was calculated and the regimen with the lowest DRS was selected as the best initial treatment for each patient. The best initial treatment of 16 patients was CPT-11, of 8 patients was O-FP, of 2 patients was Pt-2d and of 1 patient was PTX. Following that, the optimal program of sequential chemotherapy was designed according to the percentage of shared resistant genes between regimens for each patient, where two regimens with the lowest percentage were recommended to use one by one. Finally, prognostic analysis showed that differences between the optimal and practical program of sequential chemotherapy were negatively correlated to overall survival of patients. Conclusions: Based on gene-level CNVs called from ctDNA of blood samples, we determined the best initial treatment and designed the optimal program of sequential chemotherapy for advanced GCA patients, pacing a key step toward precision medicine in GCA field.

Dietary Enteromorpha Polysaccharides Supplementation Improves Breast Muscle Yield and Is Associated With Modification of mRNA Transcriptome in Broiler Chickens.

The present study evaluated the effects of dietary supplementation of Enteromorpha polysaccharides (EP) on carcass traits of broilers and potential molecular mechanisms associated with it. This study used RNA-Sequencing (RNA-Seq) to detect modification in mRNA transcriptome and the cognate biological pathways affecting the carcass traits. A total of 396 one-day-old male broilers (Arbor Acres) were randomly assigned to one of six dietary treatments containing EP at 0 (CON), 1000 (EP_1000), 2500 (EP_2500), 4000 (EP_4000), 5500 (EP_5500), and 7000 (EP_7000) mg/kg levels for a 35-d feeding trial with 6 replicates/treatment. At the end of the feeding trial, six birds (one bird from each replicate cage) were randomly selected from each treatment and slaughtered for carcass traits analysis. The results showed that the dietary supplementation of EP_7000 improved the breast muscle yield (p < 0.05). Subsequently, six breast muscle samples from CON and EP_7000 groups (three samples from each group) were randomly selected for RNA-Seq analysis. Based on the RNA-Seq results, a total of 154 differentially expressed genes (DEGs) were identified (p < 0.05). Among the DEGs, 112 genes were significantly upregulated, whereas 42 genes were significantly down-regulated by EP_7000 supplementation. Gene Ontology enrichment analysis showed that the DEGs were mainly enriched in immune-related signaling pathways, macromolecule biosynthetic, DNA-templated, RNA biosynthetic, and metabolic process (p < 0.05). Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the DEGs were enriched in signaling pathways related to viral infectious diseases and cell adhesion molecules (p < 0.05). In conclusion, dietary inclusion of EP_7000 improves the breast muscle yield, which may be involved in improving the immunity and the cell differentiation of broilers, thus promoting the muscle growth of broilers. These findings could help understand the molecular mechanisms that enhance breast muscle yield by dietary supplementation of EP in broilers.

Long Non-coding RNA MRUL Contributes to Osteosarcoma Progression Through the miR-125a-5p/FUT4 Axis.

Osteosarcoma (OS) originates in the skeletal system and has a rising global incidence. Long Non-coding RNAs (lncRNAs) are key regulators of human cancers development and progression. However, their roles in the development of OS are not well understood. This research aimed to investigate the effect of a long non-coding RNA (lncRNA), MRUL, on OS and revealed its potential molecular mechanisms. The bioinformatics analysis demonstrated that lncRNA MRUL was involved in regulating nucleic acid-templated transcription, cellular macromolecule biosynthetic process, immune response, and inflammatory response. In this work, the expression of lncRNA MRUL was detected by quantitative real-time polymerase chain reaction (qRT-RCR) in both cancer tissues and cell lines. We found that lncRNA MRUL was up-regulated in cancer tissues and cell lines. Functional experiments showed that knockdown of lncRNA MRUL inhibited OS cell proliferation, and metastasis. At the same time, we found that lncRNA MRUL interacted with miR-125a-5p to suppress FUT4 expression. Moreover, inhibition of miR-125a-5p abrogated the biological roles of lncRNA MRUL knockdown on OS cell proliferation, migration, and invasion. In conclusion, these results demonstrated that OS-upregulated lncRNA MRUL promoted cell proliferation, and metastasis via negatively regulating miR-125a-5p, and imply that lncRNA MRUL may be a potential biomarker for OS.

Overexpression of miR-340-5p Inhibits Skin Fibroblast Proliferation by Targeting Kruppel-like Factor 2.

MicroRNA (miR)-340-5p has been identified to play a key role in several cancers. However, the function of miR-340-5p in skin fibroblasts remains largely unknown. Gain of function experiments were performed by infecting normal skin fibroblast cells with a lentivirus carrying 22-bp miR-340-5p. Cell proliferation was detected by Cell Counting Kit-8 (CCK-8) assay. To uncover the mechanisms, mRNA-seq was used. Differentially expressed mRNAs were further determined by Gene Ontology and KEGG pathway analyses. The protein levels were analysed by Western blotting. A dual-luciferase reporter assay was used to detect the direct binding of miR-340-5p with the 3'UTR of Kruppel-like factor 2 (KLF2). MiR-340-5p lentivirus infection suppressed normal skin fibroblast proliferation. The mRNAseq data revealed that 41 mRNAs were differentially expressed, including 22 upregulated and 19 downregulated transcripts in the miR-340-5p overexpression group compared with those in the control group. Gene Ontology and KEGG pathway analyses revealed that miR-340-5p overexpression correlated with the macromolecule biosynthetic process, cellular macromolecule biosynthetic process, membrane, and MAPK signalling pathway. Bioinformatics analysis and luciferase reporter assays showed that miR-340-5p binds to the 3'UTR of KLF2. Forced expression of miR-340-5p decreased the expression of KLF2 in normal skin fibroblasts. Overexpression of KLF2 restored skin fibroblast proliferation in the miR-340-5p overexpression group. This study demonstrates that miR-340-5p may suppress skin fibroblast proliferation, possibly through targeting KLF2. These findings could help us understand the function of miR-340-5p in skin fibroblasts. miR-340-5p could be a therapeutic target for preventing scarring.

Association Analysis of Deep Genomic Features Extracted by Denoising Autoencoders in Breast Cancer

Artificial intelligence-based unsupervised deep learning (DL) is widely used to mine multimodal big data. However, there are few applications of this technology to cancer genomics. We aim to develop DL models to extract deep features from the breast cancer gene expression data and copy number alteration (CNA) data separately and jointly. We hypothesize that the deep features are associated with patients’ clinical characteristics and outcomes. Two unsupervised denoising autoencoders (DAs) were developed to extract deep features from TCGA (The Cancer Genome Atlas) breast cancer gene expression and CNA data separately and jointly. A heat map was used to view and cluster patients into subgroups based on these DL features. Fisher’s exact test and Pearson’ Chi-square test were applied to test the associations of patients’ groups and clinical information. Survival differences between the groups were evaluated by Kaplan–Meier (KM) curves. Associations between each of the features and patient’s overall survival were assessed using Cox’s proportional hazards (COX-PH) model and a risk score for each feature set from the different omics data sets was generated from the survival regression coefficients. The risk scores for each feature set were binarized into high- and low-risk patient groups to evaluate survival differences using KM curves. Furthermore, the risk scores were traced back to their gene level DAs weights so that the three gene lists for each of the genomic data points were generated to perform gene set enrichment analysis. Patients were clustered into two groups based on concatenated features from the gene expression and CNA data and these two groups showed different overall survival rates (p-value = 0.049) and different ER (Estrogen receptor) statuses (p-value = 0.002, OR (odds ratio) = 0.626). All the risk scores from the gene expression and CNA data and their concatenated one were significantly associated with breast cancer survival. The patients with the high-risk group were significantly associated with patients’ worse outcomes (p-values ≤ 0.0023). The concatenated risk score was enriched by the AMP-activated protein kinase (AMPK) signaling pathway, the regulation of DNA-templated transcription, the regulation of nucleic acid-templated transcription, the regulation of apoptotic process, the positive regulation of gene expression, the positive regulation of cell proliferation, heart morphogenesis, the regulation of cellular macromolecule biosynthetic process, with FDR (false discovery rate) less than 0.05. We confirmed DAs can effectively extract meaningful genomic features from genomic data and concatenating multiple data sources can improve the significance of the features associated with breast cancer patients’ clinical characteristics and outcomes.

Hub genes and gene functions associated with postmenopausal osteoporosis predicted by an integrated method.

Postmenopausal osteoporosis (PO) imposes great burden on individuals and society. This study predicted hub genes and gene functions for PO by an integration of the convergent evidence (CE) method, rank product (RP) algorithm and the combing of P-values. Using the gene expression data, genes were ranked by the CE method, RP algorithm and combing P-values, respectively. Subsequently, the top 100 genes were selected from each of the three gene lists, and then the common genes for two or three methods were denoted as informative genes of PO. A mutual information network (MIN) was constructed for the informative genes utilizing the context likelihood of relatedness algorithm. Topological centrality (degree) analysis was conducted on the MIN to investigate hub genes. Then we performed Gene Ontology (GO) enrichment analysis dependent upon the Biological Networks Gene Ontology tool (BiNGO) plugin of Cytoscape to investigate hub gene functions for PO patients. Consequently, a total of 82 informative genes were obtained by integrating the results of the three methods. There were 82 nodes and 1,741 edges in the MIN, of which 8 hub genes were identified, such as PFN1, EEF2 and S100A9. The result of GO enrichment analysis showed that 49 GO terms with P<0.001 were detected, especially the top 5 gene sets were defined as hub gene functions of PO, for instance, translational elongation, translation and cellular macromolecule biosynthetic process. In conclusion, we have predicted 8 hub genes and 5 hub gene functions associated with PO patients. The findings might help understand the molecular mechanism underlying PO.

Integrated transcriptome interactome study of oncogenes and tumor suppressor genes in breast cancer

Breast cancer is the leading cause for mortality among women worldwide. Dysregulation of oncogenes and tumor suppressor genes is the major reason for the cause of cancer. Understanding these genes will provide clues and insights about their regulatory mechanism and their interplay in cancer. In the present study, an attempt is made to compare the functional characteristics and interactions of oncogenes and tumor suppressor genes to understand their biological role. 431 breast cancer samples from seven publicly available microarray datasets were collected and analysed using GEO2R tool. The identified 416 differentially expressed genes were classified into five gene sets as oncogenes (OG), tumor suppressor genes (TSG), druggable genes, essential genes and other genes. The gene sets were subjected to various analysis such as enrichment analysis (viz., GO, Pathways, Diseases and Drugs), network analysis, calculation of mutation frequencies and Guanine-Cytosine (GC) content. From the results, it was observed that the OG were having high GC content as well as high interactions than TSG. Moreover, the OG are found to have frequent mutations than TSG. The enrichment analysis results suggest that the oncogenes are involved in positive regulation of cellular protein metabolic process, macromolecule biosynthetic process and majorly in cell cycle and focal adhesion pathway in cancer. It was also found that these oncogenes are involved in other diseases such as skin diseases and viral infections. Collagenase, paclitaxel and docetaxel are some of the drugs found to be enriched for oncogenes.

Novel insights into the molecular mechanisms underlying the beneficial effects of exercise training on pulmonary arterial hypertension.

Recent animal and clinical studies report that exercise training exerts positive influences in pulmonary arterial hypertension (PAH); however, the underlying mechanisms are largely unknown. To give insight into the molecular mechanisms of the improvement effects, we performed gene expression analysis. Three Gene Expression Omnibus (GEO) datasets were analyzed, including peripheral blood mononuclear (PBMC) gene expression profiles of exercise training in men and patients with PAH. Differentially expressed genes (DEGs) in each dataset were identified, and then, the common DEGs positively regulated by PAH and negatively regulated by exercise training, or the opposite, were further identified. Subsequently, biological processes and pathways were analyzed. A total of 7229 DEGs with logFC>0.3 and P<0.05 were identified in exercise, whereas 749 and 2207 DEGs were identified in PAH from the two datasets. After overlapping the whole DEGs from all three datasets, total 16 common DEGs were identified, including BCLAF1, SATB1 and ZFP36L2. Seven of them were up-regulated in exercise training and down-regulated in PAH, and the others were opposite. In addition, these common DEGs were mainly enriched in negative regulation of cellular process, negative regulation of biological process and negative regulation of cellular macromolecule biosynthetic process. Some genes have been implicated in the improvement of pulmonary vascular remodeling and PAH. These findings could not only improve the knowledge about the molecular mechanisms underlying the beneficial effects of exercise training on PAH, but also provide clues for further clinical and animal studies.

Microarray‑based bioinformatics analysis of the prospective target gene network of key miRNAs influenced by long non‑coding RNA PVT1 in HCC.

The long non-coding RNA (lncRNA) PVT1 plays vital roles in the tumorigenesis and development of various types of cancer. However, the potential expression profiling, functions and pathways of PVT1 in HCC remain unknown. PVT1 was knocked down in SMMC-7721 cells, and a miRNA microarray analysis was performed to detect the differentially expressed miRNAs. Twelve target prediction algorithms were used to predict the underlying targets of these differentially expressed miRNAs. Bioinformatics analysis was performed to explore the underlying functions, pathways and networks of the targeted genes. Furthermore, the relationship between PVT1 and the clinical parameters in HCC was confirmed based on the original data in the TCGA database. Among the differentially expressed miRNAs, the top two upregulated and downregulated miRNAs were selected for further analysis based on the false discovery rate (FDR), fold-change (FC) and P-values. Based on the TCGA database, PVT1 was obviously highly expressed in HCC, and a statistically higher PVT1 expression was found for sex (male), ethnicity (Asian) and pathological grade (G3+G4) compared to the control groups (P<0.05). Furthermore, Gene Ontology (GO) analysis revealed that the target genes were involved in complex cellular pathways, such as the macromolecule biosynthetic process, compound metabolic process, and transcription. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the MAPK and Wnt signaling pathways may be correlated with the regulation of the four candidate miRNAs. The results therefore provide significant information on the differentially expressed miRNAs associated with PVT1 in HCC, and we hypothesized that PVT1 may play vital roles in HCC by regulating different miRNAs or target gene expression (particularly MAPK8) via the MAPK or Wnt signaling pathways. Thus, further investigation of the molecular mechanism of PVT1 in HCC is needed.

Identification of key genes and pathways in uterine leiomyosarcoma through bioinformatics analysis.

Uterine leiomyosarcoma (uLMS) is a rare but malignant gynaecological tumour with a poor survival outcome. The present study was aimed at identifying the key genes and pathways in the development of uLMS through bioinformatics analysis. To minimize the frequency of false-positive results of the bioinformatics analysis, 3 microarrays including GSE764, GSE64763 and GSE68312 were downloaded from Gene Expression Omnibus (GEO). Differentially expressed genes (DEGs) were screened out using the online tool GEO2R. Then, Gene Ontology and Kyoto Encyclopaedia of Genes and Genomes pathway enrichment analyses were performed using the Database for Annotation, Visualization and Integrated Discovery. Finally, a protein-protein interaction (PPI) network of the DEGs was constructed using Cytoscape, and module analysis was conducted using the plug-in MCODE. A total of 95 DEGs including 21 upregulated genes and 74 downregulated genes were identified. The upregulated DEGs were annotated with 'DNA metabolic process', 'nucleobase-containing compound biosynthetic process' and 'cellular macromolecule biosynthetic process', while the downregulated DEGs were annotated with 'cellular response to chemical stimulus', 'movement of cell or subcellular component' and 'response to inorganic substances'. The results of the PPI network analysis demonstrated that matrix metallopeptidase 9, apolipoprotein E, cyclin E1 and syndecan 1 were the predominant upregulated genes in uLMS. Additionally, the genes in the main module were enriched in 'proteoglycans in cancer', 'p53 signalling pathway' and 'extracellular matrix-receptor interaction'. The key genes and pathways identified in the present study may provide valuable clues for clarifying the molecular mechanism underlying the development of uLMS and demonstrate promise for use as diagnostic markers and therapeutic targets.

Identification of key genes in non‑alcoholic fatty liver disease progression based on bioinformatics analysis.

Due to economic development and lifestyle changes, the incidence of non-alcoholic fatty liver disease (NAFLD) has gradually increased in recent years. However, the pathogenesis of NAFLD is not yet fully understood. To identify candidate genes that contribute to the development and progression of NAFLD, two microarray datasets were downloaded from the Gene Expression Omnibus database. The differentially expressed genes (DEGs) were identified and functional enrichment analyses were performed. A protein-protein interaction network was constructed and modules were extracted using the Search Tool for the Retrieval of Interacting Genes and Cytoscape. The enriched functions and pathways of the DEGs included ‘cellular macromolecule biosynthetic process’, ‘cellular response to chemical stimulus’, ‘extracellular matrix organization’, ‘metabolic pathways’, ‘insulin resistance’ and ‘forkhead box protein O1 signaling pathway’. The DEGs, including type-1 angiotensin II receptor, formin-binding protein 1-like, RNA-binding protein with serine-rich domain 1, Ras-related C3 botulinum toxin substrate 1 and polyubiquitin-C, were identified using multiple bioinformatics methods and validated in vitro with reverse transcription-quantitative polymerase chain reaction analysis. In conclusion, five hub genes were identified in the present study, and they may aid in understanding of the molecular mechanisms underlying the development and progression of NAFLD.

High-Throughput Sequencing of Small RNA Transcriptomes in Maize Kernel Identifies miRNAs Involved in Embryo and Endosperm Development.

Maize kernel development is a complex biological process that involves the temporal and spatial expression of many genes and fine gene regulation at a transcriptional and post-transcriptional level, and microRNAs (miRNAs) play vital roles during this process. To gain insight into miRNA-mediated regulation of maize kernel development, a deep-sequencing technique was used to investigate the dynamic expression of miRNAs in the embryo and endosperm at three developmental stages in B73. By miRNA transcriptomic analysis, we characterized 132 known miRNAs and six novel miRNAs in developing maize kernel, among which, 15 and 14 miRNAs were commonly differentially expressed between the embryo and endosperm at 9 days after pollination (DAP), 15 DAP and 20 DAP respectively. Conserved miRNA families such as miR159, miR160, miR166, miR390, miR319, miR528 and miR529 were highly expressed in developing embryos; miR164, miR171, miR393 and miR2118 were highly expressed in developing endosperm. Genes targeted by those highly expressed miRNAs were found to be largely related to a regulation category, including the transcription, macromolecule biosynthetic and metabolic process in the embryo as well as the vitamin biosynthetic and metabolic process in the endosperm. Quantitative reverse transcription-PCR (qRT-PCR) analysis showed that these miRNAs displayed a negative correlation with the levels of their corresponding target genes. Importantly, our findings revealed that members of the miR169 family were highly and dynamically expressed in the developing kernel, which will help to exploit new players functioning in maize kernel development.