Deregulation Of Gene Expression Research Articles

Severe Congenital Neutropenia-Associated Mutations Reveal Stage-Specific Gfi1-Dependent Checkpoints in Myeloid Development

Growth factor independent-1 (GFI1) is a zinc-finger transcription factor that is critically required for both murine and human granulopoiesis. Mutations in GFI1 zinc fingers (e.g. N382S in zinc finger 5) have previously been found in humans with severe congenital neutropenia (SCN). We noted that clinical resequencing of neutropenic patients identified a number of novel GFI1 sequence changes of unknown significance. To clarify their impact, we used viral vectors to force expression of 13 GFI1 polymorphisms/mutations in primary-murine progenitors followed by colony-forming-unit assays for differentiation. While these assays infer new potential clinical significance to some genetic alterations, we focused on the sixth zinc finger of GFI1. While Gfi1 zinc-finger 6 was previously found to be dispensable for DNA binding and repression of a high affinity GFI1 DNA-binding-site reporter, the biological impact of expressing GFI1 zinc finger 6 mutants suggests a functional role for this domain in granulopoiesis. Therefore, we generated mice with patient-derived SCN-associated mutations (zinc finger 5; N382S, zinc finger 6; K403R, and R412X) in the murine Gfi1 locus. The resulting defects in murine neutrophil production are proportional to the severity of SCN in patients with the same GFI1 mutation (ranging from mild to profound neutropenia); with neonates displaying more profound defects. Neonatal and adult Gfi1N382S/- and Gfi1R412X/- mice are neutropenic, but Gfi1K403R/- mice have normal steady-state neutrophil levels. While Gfi1-/- HSC are profoundly abnormal, SCN-associated mutant HSC are phenotypically normal; dissociating Gfi1 control of hematopoietic stem cell biology from granulopoiesis. Similar to Gfi1-/-, the Gfi1N382S/- and Gfi1R412X/- mice accumulate cells within the “granulocyte monocyte progenitor” (GMP) gate as well as those that express Sca1 (known as emergency GMP; eGMP); however, unlike Gfi1-/- mice, the serum levels of known cytokine inducers of eGMP are normal in SCN-associated mutants. Moreover, isolated progenitors from all three mutants form significantly more monocytic colonies at the expense of granulocytic colonies in methylcellulose assays. Together, these data suggest that the mutations induce a cell-intrinsic defect.We recently utilized single-cell RNA-Seq (scRNA-Seq) to identify a gene regulatory network (GRN) centered on the Gfi1 and Irf8 transcription factors that underlies the granulocytic versus monocytic lineage decision of myeloid progenitors at steady state. To pinpoint the molecular deregulation underlying Gfi1-mutant progenitors, we performed scRNA-Seq. Notably, Gfi1R412X/- induced deregulation of gene expression which increases in amplitude and complexity as myeloid differentiation proceeds; coincident with induction of Gfi1 expression. ChIP-Seq analysis reveals that most of the deregulated genes are normally bound by Gfi1. Using murine fluorescent-protein knock-in reporter alleles for Gfi1 and Irf8, we demonstrate the stage-specific regulation of the granulocyte-monocyte lineage decision, and how SCN mutations deregulate the Gfi1-Irf8 network. Finally, we developed an alternative flow-cytometric strategy that highlights individual Gfi1-dependent checkpoints in myeloid differentiation (identified by comparative analysis of Gfi1-/- versus knock-in mutants). We find that the accumulated Gfi1N382S/- GMP are unspecified (similar to Gfi1-/-); however, Gfi1R412X/- GMP exhibit only a modest reduction in the granulocytic specified population and instead display a profound block at granulocytic commitment. The latter effects can be partially explained by the protein-destabilizing effect of mutations in zinc finger 6; suggesting that there are dose-dependent roles for Gfi1 in granulopoietic specification and commitment. In summary, Gfi1 knock-in mice provide a mouse model of human SCN, and both flow-cytometric and scRNA-Seq dissection of hematopoiesis reveal myelopoietic-stage-specific deregulation to both block granulopoiesis and deregulate monocytic subsets. DisclosuresNo relevant conflicts of interest to declare.

Identification of Novel Fusion Genes and Characterization of Transcriptome Features in T-Cell Acute Lymphoblastic Leukemia

T-cell acute lymphoblastic leukemia (T-ALL) is a clonal malignancy of immature T cells, accounting for 15% of pediatric ALL and 25% of adult ALL. Significant therapeutic progress has been made in pediatric T-ALL, while the outcome remains poor in adult T-ALL. Therefore, it is important to further characterize the molecular pathogenesis of T-ALL. Recently, the next-generation sequencing approach allowed systematic identification of molecular markers in pediatric T-ALL.Here, by performing RNA-sequencing and other genomic analysis, we investigated genomic landscape in 130 T-ALL cases. 36 distinct gene fusions were identified, including 18 novel ones. Four types of gene abnormalities were identified among novel fusions. Of note, newly discovered ZBTB16-ABL1 and TRA-SALL2 fusions, and the involvement of NKX2-1, were recurrent events. Functional studies revealed the recurrent ZBTB16-ABL1 fusion as a leukemogenic driver. Among 48 genes with mutation rates>3%, 6 were newly found in T-ALL. Moreover, aberrant mRNA isoform of SLC17A9 gene was found in majority of cases with high expression of TAL1 . Although most fusions were mutually exclusive, they co-existed with mutations. These genetic abnormalities were correlated to deregulated gene expression markers in three transcriptome-based subgroups. The G1 group mainly harbored TLX1 or TLX3 high expression, and over-expression of HOXA gene family. The transcriptome landscape of G2 group was closely associated with NUP family genes-containing fusions. The transcriptional factors important for early hematopoiesis were also found highly expressed in this group. Notably, 91.7% of our ETP cases were clustered in G2 group. With regard to G3, all patients with STIL-TAL1 and LMO1/2-TRA were located in this group, as cases with variant fusions involving TAL1 or STIL did. The genes with transcriptional aberration related to STIL-TAL1 were found over-expressed in most cases of G3 group. Finally, we compared genomic landscapes between adult and pediatric T-ALL patients. The types of fusions seemed more diverse in children than in adults, and the mutation rates in the genes of signaling pathways and epigenetic factors, both of which been frequently seen in ETP-ALL, were much higher in adults than in children. Our data suggest that the two T-ALL age groups might represent distinct disease entities, which could explain, at least in part, their distinct clinical outcomes. This study shed new lights on the leukemogenesis of T-ALL. DisclosuresNo relevant conflicts of interest to declare.

Regulation of transgelin and GST-pi proteins in the tissues of hamsters exposed to sodium arsenite

Regulation of transgelin and GST-pi proteins in the tissues of hamsters exposed to sodium arsenite

Does and duration of exposure time dependent regulation of proteins in the urinary bladder of hamsters exposed to sodium arsenite

Does and duration of exposure time dependent regulation of proteins in the urinary bladder of hamsters exposed to sodium arsenite

ALS-linked FUS mutants affect the localization of U7 snRNP and replication-dependent histone gene expression in human cells

Genes encoding replication-dependent histones lack introns, and the mRNAs produced are a unique class of RNA polymerase II transcripts in eukaryotic cells that do not end in a polyadenylated tail. Mature mRNAs are thus formed by a single endonucleolytic cleavage that releases the pre-mRNA from the DNA and is the only processing event necessary. U7 snRNP is one of the key factors that determines the cleavage site within the 3ʹUTR of replication-dependent histone pre-mRNAs. We have previously showed that the FUS protein interacts with U7 snRNA/snRNP and regulates the expression of histone genes by stimulating transcription and 3ʹ end maturation. Mutations in the FUS gene first identified in patients with amyotrophic lateral sclerosis (ALS) lead to the accumulation of the FUS protein in cytoplasmic inclusions. Here, we report that mutations in FUS lead to disruption of the transcriptional activity of FUS and mislocalization of U7 snRNA/snRNP in cytoplasmic aggregates in cellular models and primary neurons. As a consequence, decreased transcriptional efficiency and aberrant 3ʹ end processing of histone pre-mRNAs were observed. This study highlights for the first time the deregulation of replication-dependent histone gene expression and its involvement in ALS.

Pyrrolizidine alkaloids cause cell cycle and DNA damage repair defects as analyzed by transcriptomics in cytochrome P450 3A4-overexpressing HepG2 clone 9 cells

Pyrrolizidine alkaloids (PAs) are a large group of highly toxic chemical compounds, which are found as cross-contaminants in numerous food products (e.g., honey), dietary supplements, herbal teas, and pharmaceutical herbal medicines. PA contaminations are responsible for serious hepatotoxicity and hepatocarcinogenesis. Health authorities have to set legal limit values to guarantee the safe consumption of plant-based nutritional and medical products without harmful health. Toxicological and chemical analytical methods are conventionally applied to determine legally permitted limit values for PAs. In the present investigation, we applied a highly sensitive transcriptomic approach to investigate the effect of low concentrations of five PAs (lasiocarpine, riddelliine, lycopsamine, echimidine, and monocrotaline) on human cytochrome P450 3A4-overexpressing HepG2 clone 9 hepatocytes. The transcriptomic profiling of deregulated gene expression indicated that the PAs disrupted important signaling pathways related to cell cycle regulation and DNA damage repair in the transfected hepatocytes, which may explain the carcinogenic PA effects. As PAs affected the expression of genes that involved in cell cycle regulation, we applied flow cytometric cell cycle analyses to verify the transcriptomic data. Interestingly, PA treatment led to an arrest in the S phase of the cell cycle, and this effect was more pronounced with more toxic PAs (i.e., lasiocarpine and riddelliine) than with the less toxic monocrotaline. Using immunofluorescence, high fractions of cells were detected with chromosome congression defects upon PA treatment, indicating mitotic failure. In conclusion, the tested PAs revealed threshold concentrations, above which crucial signaling pathways were deregulated resulting in cell damage and carcinogenesis. Cell cycle arrest and DNA damage repair point to the mutagenicity of PAs. The disturbance of chromosome congression is a novel mechanism of Pas, which may also contribute to PA-mediated carcinogenesis. Transcriptomic, cell cycle, and immunofluorescence analyses should supplement the standard techniques in toxicology to unravel the biological effects of PA exposure in liver cells as the primary target during metabolization of PAs.Graphical abstract

Effect and in silico characterization of genetic variants associated with severe spermatogenic disorders in a large Iberian cohort.

Severe spermatogenic failure (SpF) represents the most extreme manifestation of male infertility, as it decreases drastically the semen quality leading to either severe oligospermia (SO, <5 million spermatozoa/mL semen) or non-obstructive azoospermia (NOA, complete lack of spermatozoa in the ejaculate without obstructive causes). The main objective of the present study is to analyze in the Iberian population the effect of 6 single-nucleotide polymorphisms (SNPs) previously associated with NOA in Han Chinese through genome-wide association studies (GWAS) and to establish their possible functional relevance in the development of specific SpF patterns. We genotyped 674 Iberian infertile men (including 480 NOA and 194 SO patients) and 1058 matched unaffected controls for the GWAS-associated variants PRMT6-rs12097821, PEX10-rs2477686, CDC42BPA-rs3000811, IL17A-rs13206743, ABLIM1-rs7099208, and SOX5-rs10842262. Their association with SpF, SO, NOA, and different NOA phenotypes was evaluated by logistic regression models, and their functional relevance was defined by comprehensive interrogation of public resources. ABLIM1-rs7099208 was associated with SpF under both additive (OR=0.86, p=0.036) and dominant models (OR=0.78, p=0.026). The CDC42BPA-rs3000811 minor allele frequency was significantly increased in the subgroup of NOA patients showing maturation arrest (MA) of germ cells compared to the remaining NOA cases under the recessive model (OR=4.45, p=0.044). The PEX10-rs2477686 SNP was associated with a negative testicular sperm extraction (TESE) outcome under the additive model (OR=1.32, p=0.034). The analysis of functional annotations suggested that these variants affect the testis-specific expression of nearby genes and that lincRNA may play a role in SpF. Our data support the association of three previously reported NOA risk variants in Asians (ABLIM1-rs7099208, CDC42BPA-rs3000811, and PEX10-rs2477686) with different manifestations of SpF in Iberians of European descent, likely by influencing gene expression and lincRNA deregulation.

TRIM28 Expression on Dendritic Cells Prevents Excessive T Cell Priming by Silencing Endogenous Retrovirus.

Acquired immune reaction is initiated by dendritic cells (DCs), which present Ags to a few naive Ag-specific T cells. Deregulation of gene expression in DCs may alter the outcome of the immune response toward immunodeficiency and/or autoimmune diseases. Expression of TRIM28, a nuclear protein that mediates gene silencing through heterochromatin, decreased in DCs from old mice, suggesting alteration of gene regulation. Mice specifically lacking TRIM28 in DCs show increased DC population in the spleen and enhanced T cell priming toward inflammatory effector T cells, leading to acceleration and exacerbation in experimental autoimmune encephalomyelitis. TRIM28-deficient DCs were found to ectopically transcribe endogenous retrovirus (ERV) elements. Combined genome-wide analysis revealed a strong colocalization among the decreased repressive histone mark H3K9me3-transcribed ERV elements and the derepressed host genes that were related to inflammation in TRIM28-deficient DCs. This suggests that TRIM28 occupancy of ERV elements critically represses expression of proximal inflammatory genes on the genome. We propose that gene silencing through repressive histone modification by TRIM28 plays a role in maintaining the integrity of precise gene regulation in DCs, which prevents aberrant T cell priming to inflammatory effector T cells.

Assessment of INPP4B Expression Level in Acute Myeloid Leukemia Patients and Its Prognostic Significance. An Egyptian Study

Background: Acute myeloid leukemia (AML) is a complex and heterogeneous hematopoietic tissue neoplasm caused by gene mutations, chromosomal rearrangement and deregulation of gene expression. Inositol polyphosphate 4-phosphatase type II (INPP4B) is a clinically relevant factor in the phosphoinositide 3 kinase (PI3K) pathway- associated cancers, has been found to have a bad prognostic role in AML. However, the exact mechanism is still unclear. Objective: We aimed in this study to investigate the prognostic role of INPP4B overexpression in Egyptian AML patients. Patients and methods: A total of 80 patients with newly diagnosed AML were included in the study. In addition, 40 apparently healthy, age and sex matched subjects served as control group. Immunophenotyping, cytogenetic analysis and quantitative assessment of INPP4B gene transcript were performed using real time PCR. Results: INPP4B overexpression was detected in 27.5 % of newly diagnosed AML patients. There was a statistically significant decrease in the probability of achieving complete remission (CR) with shorter overall survival (OS), event free survival (EFS) and disease-free survival (DFS) in the INPP4B high expression group compared with the low expression group (p=0.003, 0.03, 0.02 and <0.001; respectively). Conclusion: We can conclude thatINPP4B overexpression is associated with poor response to therapy with poor outcome, and add prognostic value in patients with AML. INPP4B overexpression could be a valuable tool for making therapeutic decisions.

Caffeic acid prevents non-alcoholic fatty liver disease induced by a high-fat diet through gut microbiota modulation in mice

Caffeic acid (CA) is derived from many plants and may have the ability to reduce hepatic lipid accumulation. The gut microbiota produces lipopolysaccharides and further influences hepatic lipid metabolism, and thus plays an important role in the development of nonalcoholic fatty liver disease (NAFLD). However, whether the beneficial effects of CA are associated with the gut microbiota remains unclear. The present study aimed to investigate the benefits of experimental treatment with CA on the gut microbiota and metabolic functions in a mouse model of NAFLD. In this study, C57BL/6J mice received a high-fat diet (HFD) for 8 weeks and were then fed a HFD supplemented with or without CA for another 8 weeks. HFD induced obesity and increased accumulation of intrahepatic lipids, serum biochemical parameters and gene expression related to lipid metabolism. Microbiota composition was determined via 16S rRNA sequencing, and analysis revealed that HFD led to dysbiosis, accompanied by endotoxemia and low-grade inflammation. CA reverted the imbalance in the gut microbiota and related lipopolysaccharide-mediated inflammation, thus inhibiting deregulation of lipid metabolism-related gene expression. Our results support the possibility that CA can be used as a therapeutic approach for obesity-associated NAFLD via its anti-inflammatory and prebiotic integrative response.

The Non-coding MicroRNA-223 is a Promising Biomarker of Chronic Kidney Disease

Renal diseases are consecutive to a deregulation of gene expression regulated by non-coding RNAs. These non-coding RNAs were discovered at the turn of the 21st century when it was established that post-transcriptional regulation was performed through small non-coding RNAs, known as microRNAs (miRNAs). Up to 3,000 miRNAs are expressed by human cells. They are small, single-stranded nucleic acids, which trigger translational repression of mRNA by base-pairing with the 3′ untranslated region of their mRNA targets. In addition to miRNA regulation, it was also demonstrated that 60,000 long non-coding RNAs are expressed in the human cell and that they are able to regulate gene expression at all levels. The roles of these various RNA families are just beginning to be understood in the field of nephrology. In the past decade, the authors and various others have published that several miRNAs are deregulated during the onset of chronic kidney disease (CKD) and are associated with cardiovascular damage. This review focuses on miRNA-223 (miR-223) as its expression is increased in vivo in the large vessels of a mouse model of CKD, whereas it is diminished in the serum of both mice and human patients with CKD. In patients, miR-223 expression was correlated with all-cause mortality, as well as cardiovascular and renal events. Molecular clues were given by a multi-omics approach, indicating that miR-223 modulates gene regulation at all levels including mRNA expression, protein amounts, and metabolic molecule accumulation. miR-223 is thus a potential target to prevent or treat complications of CKD pathogenesis.

Histone H3 lysine 4 trimethylation in sperm is transmitted to the embryo and associated with diet-induced phenotypes in the offspring.

A father's lifestyle impacts offspring health; yet, the underlying molecular mechanisms remain elusive. We hypothesized that a diet that changes methyl donor availability will alter the sperm and embryo epigenomes to impact embryonic gene expression and development. Here, we demonstrate that a folate-deficient (FD) dietalters histone H3 lysine 4 trimethylation (H3K4me3) in sperm at developmental genes and putative enhancers. A subset of H3K4me3 alterations in sperm are retained in the pre-implantation embryo and associated with deregulated embryonic gene expression. Using a genetic mouse model in which sires have pre-existing altered H3K4me2/3 in sperm, we show that a FD diet exacerbates alterations in sperm H3K4me3 and embryonic gene expression, leading to an increase in developmental defect severity. These findings imply that paternal H3K4me3 is transmitted to the embryo and influences gene expression and development. It further suggests that epigenetic errors can accumulate in sperm to worsen offspring developmental outcomes.

Signs of Deregulated Gene Expression Are Present in Both CD14+ and CD14- PBMC From Non-Obese Men With Family History of T2DM.

AimDevelopment of type 2 diabetes (T2DM) is associated with disturbances in immune and metabolic status that may be reflected by an altered gene expression profile of peripheral blood mononuclear cells (PBMC). To reveal a potential family predisposition to these alterations, we investigated the regulation of gene expression profiles in circulating CD14+ and CD14- PBMC in fasting conditions and in response to oral glucose tolerance test (OGTT) in glucose tolerant first-degree relatives (FDR) of T2DM patients and in control subjects.Materials and MethodsThis work is based on the clinical study LIMEX (NCT03155412). Non-obese 12 non-diabetic (FDR), and 12 control men without family history of diabetes matched for age and BMI underwent OGTT. Blood samples taken before and at the end of OGTT were used for isolation of circulating CD14+ and CD14- PBMC. In these cells, mRNA levels of 94 genes related to lipid and carbohydrate metabolism, immunity, and inflammation were assessed by qPCR.ResultsIrrespectively of the group, the majority of analyzed genes had different mRNA expression in CD14+ PBMC compared to CD14- PBMC in the basal (fasting) condition. Seven genes (IRS1, TLR2, TNFα in CD14+ PBMC; ABCA1, ACOX1, ATGL, IL6 in CD14- PBMC) had different expression in control vs. FDR groups. OGTT regulated mRNA levels of nine genes selectively in CD14+ PBMC and of two genes (ABCA1, PFKL) selectively in CD14-PBMC. Differences in OGTT-induced response between FDR and controls were observed for EGR2, CCL2 in CD14+ PBMC and for ABCA1, ACOX1, DGAT2, MLCYD, and PTGS2 in CD14- PBMC.ConclusionThis study revealed a different impact of glucose challenge on gene expression in CD14+ when compared with CD14- PBMC fractions and suggested possible impact of family predisposition to T2DM on basal and OGTT-induced gene expression in these PBMC fractions. Future studies on these putative alterations of inflammation and lipid metabolism in fractionated PBMC in larger groups of subjects are warranted.

A physiological glucocorticoid rhythm is an important regulator of brown adipose tissue function

ObjectiveBrown adipose tissue (BAT) displays a strong circadian rhythm in metabolic activity, but it is unclear how this rhythm is regulated. As circulating levels of corticosterone coincide with the rhythm of triglyceride-derived fatty acid (FA) uptake by BAT, we investigated whether corticosterone regulates BAT circadian rhythm. MethodsCorticosterone levels were flattened by implanting mice with subcutaneous corticosterone-releasing pellets, resulting in constant circulating corticosterone levels. ResultsFlattened corticosterone rhythm caused a complete loss of circadian rhythm in triglyceride-derived fatty acid uptake by BAT. This effect was independent of glucocorticoid receptor expression in (brown) adipocytes and was not caused by deregulation of clock gene expression or overexposure to glucocorticoids, but rather seemed mediated by reduced sympathetic innervation of BAT. In a mouse model of hyperlipidemia and metabolic syndrome, long-term experimental flattening of corticosterone − and thus rhythm in BAT function − resulted in adiposity. ConclusionsThis study highlights that a physiological rhythm in glucocorticoids is an important regulator of BAT function and essential for the maintenance of metabolic health.

Gene Expression in Solitary Fibrous Tumors (SFTs) Correlates with Anatomic Localization and NAB2-STAT6 Gene Fusion Variants

Solitary fibrous tumors (SFTs) harbor recurrent NAB2-STAT6 gene fusions, promoting constitutional up-regulation of oncogenic early growth response 1 (EGR1)-dependent gene expression. SFTs with the most common canonical NAB2 exon 4-STAT6 exon 2 fusion variant are often located in the thorax (pleuropulmonary) and are less cellular with abundant collagen. In contrast, SFTs with NAB2 exon 6-STAT6 exon 16/17 fusion variants typically display a cellular round to ovoid cell morphology and are often located in the deep soft tissue of the retroperitoneum and intra-abdominal pelvic region or in the meninges. Here, we employed next-generation sequencing-based gene expression profiling to identify significant differences in gene expression associated with anatomic localization and NAB2-STAT6 gene fusion variants. SFTs with the NAB2 exon 4-STAT6 exon 2 fusion variant showed a transcriptional signature enriched for genes involved in DNA binding, gene transcription, and nuclear localization, whereas SFTs with the NAB2 exon 6-STAT6 exon 16/17 fusion variants were enriched for genes involved in tyrosine kinase signaling, cell proliferation, and cytoplasmic localization. Specific transcription factor binding motifs were enriched among differentially expressed genes in SFTs with different fusion variants, implicating co-transcription factors in the modification of chimeric NGFI-A binding protein 2 (NAB2)-STAT6-dependent deregulation of EGR1-dependent gene expression. In summary, this study establishes a potential molecular biologic basis for clinicopathologic differences in SFTs with distinct NAB2-STAT6 gene fusion variants.

Phototropin 1 and 2 Influence Photosynthesis, UV-C Induced Photooxidative Stress Responses, and Cell Death.

Phototropins are plasma membrane-associated photoreceptors of blue light and UV-A/B radiation. The Arabidopsis thaliana genome encodes two phototropins, PHOT1 and PHOT2, that mediate phototropism, chloroplast positioning, and stomatal opening. They are well characterized in terms of photomorphogenetic processes, but so far, little was known about their involvement in photosynthesis, oxidative stress responses, and cell death. By analyzing phot1, phot2 single, and phot1phot2 double mutants, we demonstrated that both phototropins influence the photochemical and non-photochemical reactions, photosynthetic pigments composition, stomata conductance, and water-use efficiency. After oxidative stress caused by UV-C treatment, phot1 and phot2 single and double mutants showed a significantly reduced accumulation of H2O2 and more efficient photosynthetic electron transport compared to the wild type. However, all phot mutants exhibited higher levels of cell death four days after UV-C treatment, as well as deregulated gene expression. Taken together, our results reveal that on the one hand, both phot1 and phot2 contribute to the inhibition of UV-C-induced foliar cell death, but on the other hand, they also contribute to the maintenance of foliar H2O2 levels and optimal intensity of photochemical reactions and non-photochemical quenching after an exposure to UV-C stress. Our data indicate a novel role for phototropins in the condition-dependent optimization of photosynthesis, growth, and water-use efficiency as well as oxidative stress and cell death response after UV-C exposure.

Alternative splicing in normal and pathological human placentas is correlated to genetic variants

Two major obstetric diseases, preeclampsia (PE), a pregnancy-induced endothelial dysfunction leading to hypertension and proteinuria, and intra-uterine growth-restriction (IUGR), a failure of the fetus to acquire its normal growth, are generally triggered by placental dysfunction. Many studies have evaluated gene expression deregulations in these diseases, but none has tackled systematically the role of alternative splicing. In the present study, we show that alternative splicing is an essential feature of placental diseases, affecting 1060 and 1409 genes in PE vs controls and IUGR vs controls, respectively, many of those involved in placental function. While in IUGR placentas, alternative splicing affects genes specifically related to pregnancy, in preeclamptic placentas, it impacts a mix of genes related to pregnancy and brain diseases. Also, alternative splicing variations can be detected at the individual level as sharp splicing differences between different placentas. We correlate these variations with genetic variants to define splicing Quantitative Trait Loci (sQTL) in the subset of the 48 genes the most strongly alternatively spliced in placental diseases. We show that alternative splicing is at least partly piloted by genetic variants located either in cis (52 QTL identified) or in trans (52 QTL identified). In particular, we found four chromosomal regions that impact the splicing of genes in the placenta. The present work provides a new vision of placental gene expression regulation that warrants further studies.

Colon Cancer Progression Is Reflected to Monotonic Differentiation in Gene Expression and Pathway Deregulation Facilitating Stage-specific Drug Repurposing.

Colon cancer is one of the most common cancer types and the second leading cause of death due to cancer. Many efforts have been performed towards the investigation of molecular alterations during colon cancer progression. However, the identification of stage-specific molecular markers remains a challenge. The aim of this study was to develop a novel computational methodology for the analysis of alterations in differential gene expression and pathway deregulation across colon cancer stages in order to reveal stage-specific biomarkers and reinforce drug repurposing investigation. Transcriptomic datasets of colon cancer were used to identify (a) differentially expressed genes with monotonicity in their fold changes (MEGs) and (b) perturbed pathways with ascending monotonic enrichment (MEPs) related to the number of the participating differentially expressed genes (DEGs), across the four colon cancer stages. Through an in silico drug repurposing pipeline we identified drugs that regulate the expression of MEGs and also target the resulting MEPs. Our methodology highlighted 15 MEGs and 32 candidate repurposed drugs that affect their expression. We also found 51 MEPs divided into two groups according to their rate of DEG content alteration across colon cancer stages. Focusing on the target MEPs of the highlighted repurposed drugs, we found that one of them, the neuroactive ligand-receptor interaction, was targeted by the majority of the candidate drugs. Moreover, we observed that two of the drugs (PIK-75 and troglitazone) target the majority of the resulting MEPs. These findings highlight significant genes and pathways that can be used as stage-specific biomarkers and facilitate the discovery of new potential repurposed drugs for colon cancer. We expect that the computational methodology presented can be applied in a similar way to the analysis of any progressive disease.

The Hippo pathway component Wwc2 is a key regulator of embryonic development and angiogenesis in mice

The WW-and-C2-domain-containing (WWC) protein family is involved in the regulation of cell differentiation, cell proliferation, and organ growth control. As upstream components of the Hippo signaling pathway, WWC proteins activate the Large tumor suppressor (LATS) kinase that in turn phosphorylates Yes-associated protein (YAP) and its paralog Transcriptional coactivator-with-PDZ-binding motif (TAZ) preventing their nuclear import and transcriptional activity. Inhibition of WWC expression leads to downregulation of the Hippo pathway, increased expression of YAP/TAZ target genes and enhanced organ growth. In mice, a ubiquitous Wwc1 knockout (KO) induces a mild neurological phenotype with no impact on embryogenesis or organ growth. In contrast, we could show here that ubiquitous deletion of Wwc2 in mice leads to early embryonic lethality. Wwc2 KO embryos display growth retardation, a disturbed placenta development, impaired vascularization, and finally embryonic death. A whole-transcriptome analysis of embryos lacking Wwc2 revealed a massive deregulation of gene expression with impact on cell fate determination, cell metabolism, and angiogenesis. Consequently, a perinatal, endothelial-specific Wwc2 KO in mice led to disturbed vessel formation and vascular hypersprouting in the retina. In summary, our data elucidate a novel role for Wwc2 as a key regulator in early embryonic development and sprouting angiogenesis in mice.

Romidepsin hepatocellular carcinoma suppression in mice is associated with deregulated gene expression of bone morphogenetic protein and Notch signaling pathway components.

Recently, our group showed that Romidepsin, a histone deacetylase inhibitor (HDACi), suppressed diethylnitrosamine (DEN)-induced hepatocellular carcinoma (HCC) in mice. In the present study, we investigated the effect of Romidepsin-treatment on gene expression levels of components of Bmp and Notch signaling pathways, which are both known to be aberrantly regulated in hepatocarcinogenesis. Total RNA from liver tissue samples and paraffin-embedded livers were retrieved from a recent experiment where C57BL/6 mice were treated with Romidepsin 10months after DEN challenge and sacrificed 2months later. RT qPCR was used for quantification of gene expression and immunohistochemistry for in situ protein detection. Regarding Bmp pathway, Romidepsin HCC-suppression was found to correlate significantly with Bmp2 and Bmp7 ligand up- and down-regulation, respectively. Intracellularly, Romidepsin-treated HCC mice exhibited a significant elevation of Bmp-inhibitor Smurf2 and Bmp-target gene Id3, as compared to the HCC untreated controls. Concerning Notch signaling, higher expression levels of ligands Jag1/Dll4, accompanied by a decreased expression of receptor Notch2, were identified in the Romidepsin-treated group. Τhe anti-oncogenic effect of Romidepsin, also correlated significantly with an increased expression of Hes1 target, as well as an up- and down-regulation of Klf4 and Sox9 transcription factors, respectively. Moreover, the cancer-related genes Snai2 and p21, known to be involved in many signaling pathways, including Bmp and Notch, were also found to be downregulated in Romidepsin-treated mice. Romidepsin HCC suppression is associated with gene expression deregulation of selective components of both Bmp and Notch signaling cascades.