Ribosome Pathways Research Articles

Growth, Stoichiometry, and Palatability of Suaeda salsa From Different Habitats Are Demonstrated by Differentially Expressed Proteins and Their Enriched Pathways.

Suaeda salsa (L.) Pall., a medicinal and edible plant, has green and red-violet ecotypes that exhibit different phenotypes, tastes, and growth characteristics. However, few studies have focused on these differences from the aspect of differentially expressed proteins under the conditions of different habitats in the field. In this study, two ecotypes of S. salsa from the intertidal (control) and supratidal (treatment) habitats of the Yellow River Delta were selected. A total of 30 individual leaves were mixed into six samples (three biological replicates for each) and subjected to protein extraction by using tandem mass tag-labeled quantitative proteomic technology. A total of 4771 proteins were quantitated. They included 317 differentially expressed proteins (2.0-fold change, p < 0.05), among which 143 were upregulated and the remaining 174 were downregulated. These differentially expressed proteins mainly participated in biological processes, such as response to stimulus, stress, and biotic stimulus; in molecular functions, such as methyltransferase activity, transferase activity, one-C group transfer, and tetrapyrrole binding; and in cell components, such as non-membrane-bound organelles, intracellular non-membrane-bound organelles, chromosomes, and photosystems. The differentially expressed proteins were mainly enriched in eight pathways, among which the ribosome, phenylpropanoid biosynthesis, and photosynthesis pathways had higher protein numbers than the other pathways. The upregulation of differentially expressed proteins related to the ribosome and photosynthesis increased the relative growth rate and reduced the N:P ratio of S. salsa from the supratidal habitat, thereby improving its palatability. By contrast, most of the differentially expressed proteins involved in phenylpropanoid biosynthesis were downregulated in S. salsa from the intertidal habitat. This result indicated that S. salsa from the intertidal habitat might accumulate flavonoids, lignin, and other secondary metabolites in its leaves that confer a bitter taste. However, these secondary metabolites might increase the medicinal value of S. salsa from the intertidal habitat. This work could provide a theoretical basis and data support for the sustainable and high-value utilization of medicinal and edible plants from coastal wetlands.

Ribosome Protection as a Mechanism of Lincosamide Resistance in Mycobacterium abscessus.

Mycobacterium abscessus has emerged as a successful pathogen owing to its intrinsic drug resistance. Macrolide and lincosamide antibiotics share overlapping binding sites within the ribosome and common resistance pathways. Nevertheless, while M. abscessus is initially susceptible to macrolides, they are completely resistant to the lincosamide antibiotics. Here, we have used RNA sequencing to determine the changes in gene expression in M. abscessus upon exposure to the lincosamide, clindamycin (CLY). We show that Mab_1846, encoding a putative ARE-ABCF protein, was upregulated upon exposure to macrolides and lincosamides but conferred resistance to CLY alone. A Mycobacterium smegmatis homologue of Mab_1846, Ms_5102, was similarly found to be required for CLY resistance in M. smegmatis. We demonstrate that Ms5102 mediates CLY resistance by directly interacting with the ribosomes and protecting it from CLY inhibition. Additional biochemical characterization showed that ribosome binding is not nucleotide dependent, but ATP hydrolysis is required for dissociation of Ms5102 from the ribosome as well as for its ability to confer CLY resistance. Finally, we show that in comparison to the macrolides, CLY is a potent inducer of Mab_1846 and the whiB7 regulon, such that exposure of M. abscessus to very low antibiotic concentrations induces a heightened expression of erm41, hflX, and Mab_1846, which likely function together to result in a particularly antibiotic-resistant state.

Dual RNASeq Reveals NTHi-Macrophage Transcriptomic Changes During Intracellular Persistence.

Nontypeable Haemophilus influenzae (NTHi) is a pathobiont which chronically colonises the airway of individuals with chronic respiratory disease and is associated with poor clinical outcomes. It is unclear how NTHi persists in the airway, however accumulating evidence suggests that NTHi can invade and persist within macrophages. To better understand the mechanisms of NTHi persistence within macrophages, we developed an in vitro model of NTHi intracellular persistence using human monocyte-derived macrophages (MDM). Dual RNA Sequencing was used to assess MDM and NTHi transcriptomic regulation occurring simultaneously during NTHi persistence. Analysis of the macrophage response to NTHi identified temporally regulated transcriptomic profiles, with a specific ‘core’ profile displaying conserved expression of genes across time points. Gene list enrichment analysis identified enrichment of immune responses in the core gene set, with KEGG pathway analysis revealing specific enrichment of intracellular immune response pathways. NTHi persistence was facilitated by modulation of bacterial metabolic, stress response and ribosome pathways. Levels of NTHi genes bioC, mepM and dps were differentially expressed by intracellular NTHi compared to planktonic NTHi, indicating that the transcriptomic adaption was distinct between the two different NTHi lifestyles. Overall, this study provides crucial insights into the transcriptomic adaptations facilitating NTHi persistence within macrophages. Targeting these reported pathways with novel therapeutics to reduce NTHi burden in the airway could be an effective treatment strategy given the current antimicrobial resistance crisis and lack of NTHi vaccines.

Evaluation of the Oncogene Function of GOLPH3 and Correlated Regulatory Network in Lung Adenocarcinoma.

BackgroundGolgi phosphoprotein 3 (GOLPH3) is an oncoprotein localized in the Golgi apparatus. Abnormal GOLPH3 expression is potentially related to carcinogenesis. However, the potential biological regulation network of GOLPH3 in lung adenocarcinoma (LUAD) remains to be determined.MethodsExpression of GOLPH3 was identified in LUAD via TIMER, Oncomine, Lung Cancer Explorer (LCE), Human Protein Atlas (HPA), and UALCAN database. Survival analysis was performed using the Kaplan–Meier plotter. GOLPH3 alterations were analyzed through cBioPortal. LinkedOmics was used to perform functional analysis and predict interacted targets. The protein–protein interaction network was constructed by GeneMANIA. In addition, candidate miRNAs and lncRNAs targeting GOLPH3 were generated to construct competing endogenous RNA (ceRNA) network, and survival analysis of ceRNA was performed using LnCeVar. The mRNA or protein expression of TUG1, miR-142-5p, and GOLPH3 in Beas-2B and LUAD cells was verified using qPCR or Western blotting. CCK-8 assay, wound healing assay, and transwell assay were used to detect the ability of cell proliferation, migration, and invasion.ResultsOverexpression of GOLPH3 was identified in LUAD. UALCAN analysis showed that upregulated GOLPH3 was linked to different pathological features of LUAD patients. Importantly, high GOLPH3 expression indicated a negative correlation with the first progression (FP) in LUAD patients. GOLPH3 alterations were also found. Moreover, co-expressed genes with GOLPH3 were analyzed; and they were involved in ribosome and oxidative phosphorylation pathways. Functional network analysis indicated GOLPH3 regulated T-cell receptor signaling pathway and interferon signaling pathway with kinase and transcription factor targets. Notably, TUG1/miR-142-5p/GOLPH3 affected overall survival of LUAD patients. GOLPH3 expression was decreased in the cells with overexpression of miR-142-5p and TUG1 knockdown. GOLPH3 reduction inhibited cell proliferation, migration, and invasion.ConclusionsUpregulation of GOLPH3 has a positive correlation with clinicopathological subtypes and poor FP in LUAD. GOLPH3 promoted LUAD progression. Moreover, TUG1 may act as ceRNA to regulate GOLPH3 expression by competitive binding miR-142-5p.

Exploration of the mechanism by which icariin modulates hippocampal neurogenesis in a rat model of depression

Icariin (ICA) has a significant capacity to protect against depression and hippocampal injury, but it cannot effectively cross the blood-brain barrier and accumulate in the brain. Therefore, the mechanism by which ICA protects against hippocampal injury in depression remains unclear. In this study, we performed proteomics analysis of cerebrospinal fluid to investigate the mechanism by which ICA prevents dysfunctional hippocampal neurogenesis in depression. A rat model of depression was established through exposure to chronic unpredictable mild stress for 6 weeks, after which 120 mg/kg ICA was administered subcutaneously every day. The results showed that ICA alleviated depressive symptoms, learning and memory dysfunction, dysfunctional neurogenesis, and neuronal loss in the dentate gyrus of rats with depression. Neural stem cells from rat embryonic hippocampi were cultured in media containing 20% cerebrospinal fluid from each group of rats and then treated with 100 μM corticosterone. The addition of cerebrospinal fluid from rats treated with ICA largely prevented the corticosterone-mediated inhibition of neuronal proliferation and differentiation. Fifty-two differentially expressed proteins regulated by chronic unpredictable mild stress and ICA were identified through proteomics analysis of cerebrospinal fluid. These proteins were mainly involved in the ribosome, PI3K-Akt signaling, and interleukin-17 signaling pathways. Parallel reaction monitoring mass spectrometry showed that Rps4x, Rps12, Rps14, Rps19, Hsp90b1, and Hsp90aa1 were up-regulated by chronic unpredictable mild stress and down-regulated by ICA. In contrast, HtrA1 was down-regulated by chronic unpredictable mild stress and up-regulated by ICA. These findings suggest that ICA can prevent depression and dysfunctional hippocampal neurogenesis through regulating the expression of certain proteins found in the cerebrospinal fluid. The study was approved by the Experimental Animal Ethics Committee of Guangzhou University of Chinese Medicine of China in March 2017.

RETRACTED ARTICLE: BrRNE cleaves RNA in chloroplasts, regulating retrograde signals in Brassica rapa L. ssp. pekinensis

Brassica rapa RNE participates in the processing of polycistronic precursor transcripts into mature monocistronic mRNAs in plastids, thereby sending strong retrograde signals. Leaf color is one of the most important agronomic traits for Chinese cabbage. Not only is it closely linked to photosynthesis, thereby affecting plant growth, but it also influences consumer preference in the marketplace. A pale-green mutant rne was produced by EMS mutagenesis of Chinese cabbage inbred line A03. Chlorophyll content, photosynthetic rate, actual quantum efficiency (φPSII), and maximum quantum efficiency (Fv/Fm) of photosystem II (PSII) were all reduced in rne plants. Genetic analysis indicated that the pale-green trait was controlled by a pair of recessive alleles. Using mixed pool sequencing of F2 individuals derived from an rne × wild-type cross, we identified the essential gene Brassica rapa RNase E (BrRNE), which is responsible for chloroplast development. BrRNE cleaves polycistronic RNA in Chinese cabbage A03 plastids, but rne plants are defective in RNA processing and show reduced translation levels of the seven plastid genes, BrpsaB, BrpsaA, BrpsbA, BrpsbD, BrpsbB, BrpetA, and Brycf4. Abnormal RNA processing in the plastids sends retrograde signals that markedly regulate the expression of nuclear genes, upregulating genes that participate in ribosome and DNA replication pathways and repressing photosynthesis-associated nuclear genes (PhANGs). Our study reveals a new regulatory mechanism by which plastid RNA cleavage influences plastid development and leaf color, sending retrograde signals that affect the expression of nuclear genes in Brassica.

Response of gene regulatory networks after infection of H3N2 virus.

Viral infection is a complicated dynamic process, in which viruses intrude into cells to duplicate themselves and trigger succeeding biological processes regulated by genes. It may lead to a serious disaster to human's health. A scheme is proposed to monitor the response of cells after being infected by viruses. Co-expression levels of genes measured at successive time points form a gene expression profile sequence, which is mapped to a temporal gene regulatory network. The fission and fusion of the communities of the networks are used to find the active parts. We investigated an experiment of injection of flu viruses into a total of 17 healthy volunteers, which develop into an infected group and a survival group. The survival group is much more chaotic, i.e. there occur complicated fissions and fusions of communities over the whole network. For the infected group, the most active part of the regulatory network forms a single community, but it is included in one of the large communities and completely conservative in the survival group. There are a total of six and seven genes in the active structure that take part in the Parkinson's disease and the ribosome pathways, respectively. Actually, a total of 30 genes (covering [Formula: see text]) of the genes in the active structure participate in the neuro-degeneration and its related pathways. This scheme can be extended straightforwardly to extract characteristics of trajectories of complex systems.

Screening Candidate Genes Regulating Placental Development from Trophoblast Transcriptome at Early Pregnancy in Dazu Black Goats (Capra hircus).

Simple SummaryThe trophoblast is an original placental tissue whose normal proliferation, differentiation, migration, adhesion, and angiopoiesis are essential for placenta formation and fetal survival during early pregnancy. However, the key genes and molecular mechanisms involved in placenta development in goats are unknown. Herein, the morphology and histological structures of trophoblast tissues from day 20 to 30 of pregnancy were determined. RNA-sequencing was used to screen potential functional genes in common highly expressed and differentially expressed genes. RAP1 signaling pathway was used as the contact center and coordinated with other pathways to regulate placenta development. This study could provide insights into the molecular mechanisms underlying ruminant placentation.This study explored the trophoblast transcriptome to understand potential functional genes involved in early placental development in goats and their enriched signaling pathways. Trophoblast samples were collected from nine Dazu Black goats on days 20, 25, and 30 of pregnancy (D20, D25, and D30). As the pregnancy progressed, the morphology and histological structures showed significant growth, adhesion, and angiogenesis. A total of 23,253 commonly expressed genes (CEGs) and 4439 differently expressed genes (DEGs) were detected by RNA sequencing. The common highly expressed genes (ChEGs) (the top 100 CEGs) with the highest FPKM percentage (29.9%) of all CEGs were annotated to the ribosome pathway and maintain pregnancy. DEGs were abundant in D30 vs. D20 (3715 DEGs). Besides, the DEGs were associated with the inhibition of oxidative phosphorylation and activation of PI3K-Akt, focal adhesion, ECM–receptor interaction, Rap1, and CAM signaling pathways. The RAP1 may be a central pathway since it coordinates with others to regulate the cell proliferation, invasion, migration, and fusion of trophoblasts. qRT-PCR and Western blot analysis confirmed the transcriptional expression in IGF1, VEGFC, RAPGEF3, PIK3CA, AKT3, ITGB3, ITGA11, SPP1, NOS1, and ATP6V0B genes and protein levels in VEGF, RAPGEF3, and Akt. This is the first study of transcriptome profiling in goat placenta and provides diverse genetic resources for further research on placenta development.

Oncogene-regulated release of extracellular vesicles.

Oncogenes can alter metabolism by changing the balance between anabolic and catabolic processes. However, how oncogenes regulate tumor cell biomass remains poorly understood. Using isogenic MCF10A cells transformed with nine different oncogenes, we show that specific oncogenes reduce the biomass of cancer cells by promoting extracellular vesicle (EV) release. While MYC and AURKB elicited the highest number of EVs, each oncogene selectively altered the protein composition of released EVs. Likewise, oncogenes alter secreted miRNAs. MYC-overexpressing cells require ceramide, whereas AURKB requires ESCRT to release high levels of EVs. We identify an inverse relationship between MYC upregulation and activation of the RAS/MEK/ERK signaling pathway for regulating EV release in some tumor cells. Finally, lysosome genes and activity are downregulated in the context of MYC and AURKB, suggesting that cellular contents, instead of being degraded, were released via EVs. Thus, oncogene-mediated biomass regulation via differential EV release is a new metabolic phenotype.

Niche Regulation of Residual Hematopoietic Stem Cell Protein Synthesis By Extracellular Vesicle (EV) Trafficking in the Acute Myeloid Leukemia (AML) Microenvironment

Work by several groups has shown that the presence of Acute Myeloid Leukemia (AML) deregulates critical functions of the bone marrow microenvironment, including hematopoiesis, in part through indirect stroma-secreted factors. We recently reported that Extracellular Vesicles (EV) released from AML blasts directly suppress the clonogenicity of hematopoietic progenitors via the action of miRNA-150 and miR-155 on cMyb, a transcription factor highly expressed in differentiating progenitors. The fate of long-term hematopoietic stem cells (LT- HSC) in the AML niche has not been widely studied. Using a xenograft model of human AML (Molm-14, U937 and HL-60 cell lines), we first ascertained the in vivo uptake of AML-EVs and found different internalization efficiencies among hematopoietic stem and progenitor cell (HSPC) populations, including LT-HSC (CD150+CD48−KSL) by imaging and flow cytometry. Live-cell microscopy studies of LT-HSCs revealed AML-EV binding and uptake within minutes following in vitro exposure. To determine the functional implications of EV trafficking, we next evaluated the relative frequency of the hematopoietic populations in low chimerism xenografts and found a consistent increase in the LT-HSC population, but not multi-potent progenitors, suggestingthe proportional accumulation of LT-HSC population caused by mobilization and lack of expansion and differentiation of other hematopoietic cells. To gain insight into how AML-EVs may regulate cell fate and function following internalization, we performed a proteomic profiling of HSPC cells exposed to EVs from two cell lines representing different AML subtypes. Bioinformatics analysis found significant enrichment for the ribosomal biogenesis pathways and negative cell cycle regulators in the EV-exposed cells. Because LT-HSC have been shown to be particularly susceptible to impairment of ribosome biogenesis, we wished to determine if AML-EVs modulate the LT-HSC fate by suppressing protein synthesis. We therefore injected AML-EVs into the femurs of both immuno-competent and immuno-compromised mice and evaluated the cellular protein synthesis by quantifying O-Propargyl-Puromycin (OPP) incorporation. Results show that AML-EVs significantly suppress the protein synthesis in the LT-HSC, but not in the HSPC pool population. The mTOR signaling pathway is known to be greatly involved in ribosome biogenesis via phosphorylation of the ribosomal protein S6 (S6RP) which in turn regulates several repressors of rRNA and ribosomal proteins. Consistent with the notion of translational suppression, we found that AML-EVs reduced the phosphorylation of S6RP in LT-HSC. In correlation with the observation that ribosomal stress induction via suppression of ribosomal proteins we found upregulation of P53 and its transcriptional target p21, as well as a gain in quiescence (G0) in the LT-HSC population. To address the functional fate of deregulated protein synthesis and quiescence, we transplanted KSL cells purified from male mice engrafted with AML cells into irradiated female recipient mice and measured the donor chimerism via qPCR for Sex-determining Region Y (SRY). Early repopulation capacities between cohorts of mice transplanted with KSL from AML xenograft versus those from control non-engrafted mice are similar (with long-term evaluation ongoing) in the host non-leukemic microenvironment. This suggesting that hematopoietic impairment by AML-EVs is reversible upon transplantation to a different host environment. Collectively, our results provide the first evidence that AML-EV impair LT-HSC via suppressing of ribosomal biogenesis. Understanding the regulation of LT-HSC by AML-EV will help reduce the morbidity and mortality from hematopoietic suppression in patients with leukemia. DisclosuresNo relevant conflicts of interest to declare.

RUNX3 Promotes the Development of MDS/MPN Overlap Syndrome Via Enhancing Expression of Myc in the Absence of Tet2

The RUNX transcription factors have a critical role in normal and malignant hematopoiesis including myeloproliferative neoplasms (MPN) and myelodysplastic syndromes (MDS). RUNX3 is expressed in various hematopoietic cells, and the reduced expression of RUNX3 due to promoter DNA hyper-methylation is seen in patients with acute lymphoid leukemia, indicating a tumor suppressive role of RUNX3. The loss of Runx3 promotes the development of MPN-like disease in mice. However, loss-of-function mutations and/or a deletion of RUNX3 are rarely observed in patients with myeloid malignancies. In fact, given that enhanced levels of RUNX3 expression are found in a part of patients with AML and MDS, RUNX3 may play an oncogenic role for myeloid malignancies.To determine how RUNX3 over-expression promotes the development of myeloid malignancies, we generated a mouse model of RUNX3 -induced MDS/MPN by utilizing Tet2 -deficient hematopoietic stem cells (HSCs). Since TET2 loss-of-function mutations initiate the transformation of HSCs, we harvested bone marrow CD150+CD34-LSK HSCs from tamoxifen-treated Cre-ERT2 (WT) and Cre-ERT2;Tet2Δ/Δ (Tet2 KO) mice, and transduced expression of 3xFLAG-tagged RUNX3 by using a retrovirus vector in vitro. Then, we transplanted them into lethally-irradiated Ly5.1+ recipient mice together with wild-type competitor cells. We observed that Tet2 KO mice died dominantly due to MPN-like disease, whereas RUNX3 -expressing Tet2 wild-type (RUNX3/WT) HSCs neither competed wild-type bone marrow cells nor developed hematological malignancies in mice by 12 months post transplantation. In contrast, RUNX3 -expressing Tet2 KO (RUNX3/Tet2 KO) HSCs preferentially generated myeloid cells, but not lymphoid cells, and subsequently competed wild-type cells in the PB and the BM. We found that 7 out of 18 RUNX3/Tet2 KO mice developed lethal MDS/MPN diseases despite a longer latency. At the time of sacrifice, RUNX3-Tet2 KO mice showed severe pancytopenia accompanied with dyplastic blood cells including pseudo-Pelger neutrophils and micro megakaryocytes. In addition, these moribund mice significantly enlarged spleen due to the expansion of myeloid cells. FACS analysis demonstrated that the expansion of LSK hematopoietic stem/progenitor cells (HSPCs) in the BM and the spleen, consistent with the features of extramedullary hematopoiesis. Thus, RUNX3/Tet2 KO mice recapitulated the phenotypic features of human MDS/MPN overlap syndrome, indicating that RUNX3 over-expression is prerequisite to the development of MDS/MPN in the absence of Tet2, although it has an adverse effect on the function of Tet2 -present HSCs, reminiscent with a tumor suppressive function of RUNX3 as seen in Runx3 KO mice.To understand the underlying molecular mechanism in the pathogenesis of RUNX3 -induced MDS/MPN, we performed gene expression analysis in HSPCs isolated from WT, RUNX3/WT, Tet2 KO and RUNX3/Tet2 KO mice at pre-disease and MDS/MPN stages. Gene Set Enrichment Analysis (GSEA) demonstrated that RUNX3/Tet2 KO MDS/MPN cells significantly enhanced expression of c-Myc target genes, compared to either WT cells or RUNX3/Tet2 KO cells at the pre-disease stage. While RUNX3/Tet2 KO cells showed negative enrichments of differentiation-related pathway genes, compared to WT cells, we found that those MDS/MPN cells enhanced expression of cell cycle, ribosome, and mitochondria-related biological pathways, in which c-Myc may regulate target genes and contribute to the transformation. In order to determine whether RUNX3 directly activates expression of c-Myc, we performed ChIP-sequencing by using an anti-FLAG antibody in RUNX3/Tet2 KO bone marrow cells, and found that c-Myc 3' distal enhancer regions showed conserved binding sites for Runx transcription factors. In addition, we found that RUNX3 -expressing human AML cells showed higher sensitivity to JQ1, a BRD4 inhibitor, following the significant reduction of both RUNX3 and C-MYC expression. The growth inhibition by JQ1 was rescued by ectopic expression of RUNX3 .In conclusion, RUNX3 and c-Myc collaborated to activate the function of oncogenic enhancers, in which Tet2 deficiency may alter the epigenetic status for allowing the bindings of these transcription factors, to an extent not seen in RUNX3-expressing alone. We demonstrated that RUNX3 functions as either an oncogene or a tumor suppressor for myeloid malignancies in the context dependent manner. DisclosuresHarada:Celgene: Research Funding; NIPPON SHINYAKU CO.: Speakers Bureau; NOVARTIS: Research Funding.

Growth, Immunity, and Transcriptome Response to Dietary Wnt/β-Catenin Pathway Activator TWS119 in Penaeus vannamei

Wnt/β-catenin signalling plays an essential role in the immunity of Penaeus vannamei. In this study, the effects of dietary Wnt/β-catenin pathway activator TWS119 on the growth, immunity, and transcriptome response in P. vannamei were investigated. Penaeus vannamei were fed diets with added TWS119 at doses of 0 (T0), 0.25 (T0.25), 1 (T1), 4 (T4), 16 (T16), or 64 mg·kg−1 (T64), respectively. LvGSK3β activity was effectively inhibited in P. vannamei given TWS119. The growth of P. vannamei in the T16 group was significantly improved when compared with the control group. After Vibrio parahaemolyticus infection, the survival rates (SRs) of P. vannamei in all experimental groups except the T64 group were significantly higher than in the T0 group. Compared with the control group, the immune enzymes’ activities in the serum of P. vannamei increased in all the experimental groups, while the malondialdehyde (MDA) contents decreased. Transcriptome analysis identified 5,073 differentially expressed genes (DEGs) for P. vannamei in the T0 and T16 groups. Most of the DEGs are involved in the ribosome pathway, endocytosis, glycerophospholipid metabolism, Wnt signalling, and FoxO =signalling pathways. The majority of the DEGs were from the ribosome pathway, which is also the most significantly enriched pathway. The study confirmed that the growth and immunity status of P. vannamei could improve by increasing dietary TWS119, which probably regulates the activity of the Wnt/β-catenin pathway and may be closely related to ribosome function and energy metabolism.

Identification of differentially abundant mRNA transcripts and autocrine/paracrine factors in oocytes and follicle cells of mud crabs

The late vitellogenic stage of the mud crab is characterized by large and obvious follicle cells as well as an enlarged oocyte nucleus and a prominent germinal vesicle (GV). The aim of this study was evaluation of functions of oocytes and follicle cells during meiosis as well as at identifying associated ovarian autocrine/paracrine factors using comparative transcriptomics. The results from the KEGG pathway analysis indicated DNA replication, nucleotide excision repair, spliceosome and the ribosome pathways were highly associated with oocyte maturation across both transcriptomes. In addition, there was a larger abundance of mRNA transcripts for cell cycle-related genes in the oocyte, as well as cyclin A, cyclin B and CKS1B in the GV than at the time of germinal vesicle breakdown (GVBD). These findings indicate these cell cycle-related genes might be involved in GVBD induction. Results when there was localization of ligands and the respective receptors of VEGF, TGFβ propeptide and BMP9/10 indicated these proteins might be autocrine/paracrine factors. Results from functional analysis of VEGF, TGFβ propeptide and BMP9/10 in oocyte maturation using RNA interference revealed that these proteins might be involved in oocyte maturation by regulating cyclin abundance. This is the first study on the functions of VEGF in oocyte maturation in invertebrates.

Blood Transcriptome Profiling Reveals Diet-Induced Changes in Metabolic Pathway Gene Expression

ObjectivesThis study explored the effects of diet-induced (almonds vs. crackers) changes on blood transcriptome profiles of young adults. MethodsYoung adults (age: 18–22 years) were randomly assigned to consume either almonds (2 oz./d, n = 13) or an isocaloric control snack of graham crackers (325 kcal/d, n = 10) daily for 4 weeks. Blood samples were collected at baseline and 4 weeks after intervention. Total leukocyte RNA was extracted and sequenced. Gene expression profiling was carried out using a 3′ Tag-RNA-Seq protocol. Barcoded sequencing libraries were prepared using the QuantSeq FWD kit for multiplexed sequencing. Data were preprocessed, STAR aligned, and count tables generated using the QuantSeq FWD-UMI pipeline. Differential expression (DE) analysis was conducted on the time x diet model using the limma-voom packages in R. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis on the DE p-values was conducted using the Wilcoxon rank-sum test in R. P-values were adjusted for false discovery rate correction (FDR). ResultsOut of 13,018 filtered genes, 69 were differentially expressed (FDR < 0.1). Glutamic-oxaloacetic transaminase 2 (GOT2), diacylglycerol kinase alpha (DGKA), and glutaryl-CoA dehydrogenase (GCDH) genes were upregulated with almond consumption. GOT2 plays a role in amino acid metabolism and the urea and tricarboxylic acid cycles, DGKA is involved in lipid metabolism, and GCDH is involved in lysine degradation and tryptophan metabolism. Cracker consumption resulted in greater upregulation of TGF-beta activated kinase 1 gene (MAP3K7) binding protein 3 (TAB3) gene which is involved in the NF-kappaB signal transduction pathway and a greater downregulation in the immunoglobulin superfamily member 8 (IGSF8) gene. Enrichment analyses indicate gene annotations to 341 KEGG pathways. Thermogeneic and ribosomal pathways were significantly enriched (FDR < 0.1). Pathways related to oxidative phosphorylation, sphingolipid signaling, and tryptophan, propanoate, and starch and sucrose metabolism were also differentially enriched (P < 0.05, FDR < 0.3). ConclusionsThese findings indicate that diet (almond vs cracker) potentially alters metabolism through changes in gene transcription. The implications of these findings and associations with health and disease outcomes need to be investigated further. Funding SourcesNIH

Lithium increases mitochondrial respiration in iPSC-derived neural precursor cells from lithium responders

Lithium (Li), valproate (VPA) and lamotrigine (LTG) are commonly used to treat bipolar disorder (BD). While their clinical efficacy is well established, the mechanisms of action at the molecular level are still incompletely understood. Here we investigated the molecular effects of Li, LTG and VPA treatment in induced pluripotent stem cell (iPSC)-derived neural precursor cells (NPCs) generated from 3 healthy controls (CTRL), 3 affective disorder Li responsive patients (Li-R) and 3 Li non-treated patients (Li-N) after 6 h and 1 week of exposure. Differential expression (DE) analysis after 6 h of treatment revealed a transcriptional signature that was associated with all three drugs and most significantly enriched for ribosome and oxidative phosphorylation (OXPHOS) pathways. In addition to the shared DE genes, we found that Li exposure was associated with 554 genes uniquely regulated in Li-R NPCs and enriched for spliceosome, OXPHOS and thermogenesis pathways. In-depth analysis of the treatment-associated transcripts uncovered a significant decrease in intron retention rate, suggesting that the beneficial influence of these drugs might partly be related to splicing. We examined the mitochondrial respiratory function of the NPCs by exploring the drugs’ effects on oxygen consumption rate (OCR) and glycolytic rate (ECAR). Li improved OCR levels only in Li-R NPCs by enhancing maximal respiration and reserve capacity, while VPA enhanced maximal respiration and reserve capacity in Li-N NPCs. Overall, our findings further support the involvement of mitochondrial functions in the molecular mechanisms of mood stabilizers and suggest novel mechanisms related to the spliceosome, which warrant further investigation.

The important role of MDM2, RPL5, and TP53 in mycophenolic acid-induced cleft lip and palate.

Mycophenolate embryopathy (MPE) is a mycophenolic acid (MPA)-induced congenital malformation with distinctive symptoms. Cleft lip/palate (CLP) is one of the most common symptoms of MPE. The aim of this study was to screen and verify hub genes involved in MPA-induced CLP and to explore the potential molecular mechanisms underlying MPE.Overlapping genes related to MPA and CLP were obtained from the GeneCards database. These genes were further analyzed via bioinformatics. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis results were visualized with the Cytoscape ClueGO plug-in. Gene protein-protein interaction (PPI) networks were constructed based on data obtained from the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database.Overall, 58 genes related to MPA and CLP were identified. The genes most relevant to MPA-induced CLP included ABCB1, COL1A1, Rac1, TGFβ1, EDN1, and TP53, as well as the TP53-associated genes MDM2 and RPL5. GO analysis demonstrated gene enrichment regarding such terms as ear, mesenchymal, striated muscle, and ureteric development. KEGG analysis demonstrated gene enrichment in such pathways as the HIF-1 signaling pathway, glycosylphosphatidylinositol-anchor biosynthesis, the TNF signaling pathway, and hematopoietic stem cell development.Bioinformatic analysis was performed on the genes currently known to be associated with MPA-induced CLP pathogenesis. MPA-induced CLP is mediated by multiple ribosome stress related genes and pathways. MDM2, RPL5 and TP53 could be the main contributor in this pathogenesis, along with several other genes. ABCB1 polymorphism could be related to the probability of MPA-induced CLP.

Transcriptional Responses of Fusarium graminearum Interacted with Soybean to Cause Root Rot.

Fusarium graminearum is the most devastating pathogen of Fusarium head blight of cereals, stalk and ear of maize, and it has recently become a potential threat for soybean as maize-soybean strip relay intercropping is widely practiced in China. To elucidate the pathogenesis mechanism of F. graminearum on intercropped soybean which causes root rot, transcriptional profiling of F. graminearum at 12, 24, and 48 h post-inoculation (hpi) on soybean hypocotyl tissues was conducted. In total, 2313 differentially expressed genes (DEGs) of F. graminearum were annotated by both KEGG pathway and Gene Ontology (GO) analysis. Among them, 128 DEGs were commonly expressed at three inoculation time points while the maximum DEGs were induced at 24 hpi. In addition, DEGs were also rich in carbon metabolism, ribosome and peroxisome pathways which might contribute to carbon source utilization, sexual reproduction, virulence and survival of F. graminearum when infected on soybean. Hence, this study will provide some basis for the deep understanding the pathogenesis mechanism of F. graminearum on different hosts and its effective control in maize-soybean strip relay intercropping systems.

Proteomics analysis reveals that the proto-oncogene eIF-5A indirectly influences the growth, invasion and replication of Toxoplasma gondii tachyzoite

BackgroundThe proliferative stage (tachyzoite) of Toxoplasma gondii (T. gondii) is critical for its transmission and pathogenesis, and a proto-oncogene eukaryotic translation initiation factor (eIF-5A) plays an important role in various cellular processes such as cell multiplication.MethodsWe performed a proteomic study to evaluate the specific roles of eIF-5A involved in invasion and replication of T. gondii, and both in vivo and in vitro trials using eIF-5A-interfered and wild tachyzoites were performed to verify the proteomic results.ResultsThe results of our study showed that T. gondii eIF-5A affected tachyzoite growth and also participated in the synthesis of proteins through regulation of both ribosomal and splicing pathways. Inhibition of eIF-5A in T. gondii resulted in the downregulated expression of soluble adhesions, such as microneme protein 1 (MIC1) and MIC4, which in turn decreased the parasite population that adhered to the surface of host cells. The reduced attachment, combined with lower expression of some rhoptry proteins (ROPs) and dense granule antigens (GRAs) involved in different stages of T. gondii invasion such as ROP4 and GRA3, ultimately reduce the invasion efficiency. These processes regulated by eIF-5A eventually affect the replication of tachyzoites.ConclusionsOur findings showed that eIF-5A influenced tachyzoite survival and was also involved in the process of parasite invasion and replication. These results will provide new clues for further development of targeted drugs to control T. gondii infection.Graphical abstract

Transcriptome-wide association study uncovers the role of essential genes in anthracycline-induced cardiotoxicity

Anthracyclines are highly effective chemotherapeutic agents; however, their clinical utility is limited by severe anthracycline-induced cardiotoxicity (ACT). Genome-wide association studies (GWAS) have uncovered several genetic variants associated with ACT, but the impact of these findings requires further elucidation. We conducted a transcriptome-wide association study (TWAS) using our previous GWAS summary statistics (n = 280 patients) to identify gene expression-related associations with ACT. We identified a genetic association between decreased expression of GDF5 and ACT (Z-score = −4.30, P = 1.70 × 10−5), which was replicated in an independent cohort (n = 845 patients, P = 3.54 × 10−3). Additionally, cell viability of GDF5-silenced human cardiac myocytes was significantly decreased in response to anthracycline treatment. Subsequent gene set enrichment and pathway analyses of the TWAS data revealed that genes essential for survival, cardioprotection and response to anthracyclines, as well as genes involved in ribosomal, spliceosomal and cardiomyopathy pathways are important for the development of ACT.

Comparative Transcriptomic Analysis of Spermatozoa From High- and Low-Fertile Crossbred Bulls: Implications for Fertility Prediction.

Crossbred bulls produced by crossing Bos taurus and Bos indicus suffer with high incidence of infertility/subfertility problems; however, the etiology remains poorly understood. The uncertain predictability and the inability of semen evaluation techniques to maintain constant correlation with fertility demand for alternate methods for bull fertility prediction. Therefore, in this study, the global differential gene expression between high- and low-fertile crossbred bull sperm was assessed using a high-throughput RNA sequencing technique with the aim to identify transcripts associated with crossbred bull fertility. Crossbred bull sperm contained transcripts for 13,563 genes, in which 2,093 were unique to high-fertile and 5,454 were unique to low-fertile bulls. After normalization of data, a total of 776 transcripts were detected, in which 84 and 168 transcripts were unique to high-fertile and low-fertile bulls, respectively. A total of 176 transcripts were upregulated (fold change > 1) and 209 were downregulated (<1) in low-fertile bulls. Gene ontology analysis identified that the sperm transcripts involved in the oxidative phosphorylation pathway and biological process such as multicellular organism development, spermatogenesis, and in utero embryonic development were downregulated in low-fertile crossbred bull sperm. Sperm transcripts upregulated and unique to low-fertile bulls were majorly involved in translation (biological process) and ribosomal pathway. With the use of RT-qPCR, selected sperm transcripts (n = 12) were validated in crossbred bulls (n = 12) with different fertility ratings and found that the transcriptional abundance of ZNF706, CRISP2, TNP2, and TNP1 genes was significantly (p < 0.05) lower in low-fertile bulls than high-fertile bulls and was positively (p < 0.05) correlated with conception rate. It is inferred that impaired oxidative phosphorylation could be the predominant reason for low fertility in crossbred bulls and that transcriptional abundance of ZNF706, CRISP2, TNP2, and TNP1 genes could serve as potential biomarkers for fertility in crossbred bulls.