A Comprehensive Genomic Approach Using Gain of Function Cell Models, Patient Specimens and ChIP-on-CHIP Technologies Identifies PLZF-RARα Target Genes with Potential Roles in t(11;17) Associated APL.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3168-3168
Author(s):  
Jonathan D. Licht ◽  
Kim L. Rice ◽  
Itsaso Hormaeche ◽  
Julia Meyer ◽  
Ken I. Mills ◽  
...  
Keyword(s):  
Cell Growth ◽  
Target Genes ◽  
Expression Profiles ◽  
Defense Responses ◽  
Myeloid Differentiation ◽  
Small Subset ◽  
U937 Cells ◽  
Gain Of Function ◽  
Cell Functions ◽  
On Chip

Abstract The t(11;17)(q23;q21) translocation involves the production of reciprocal fusion proteins PLZF-RARα and RARα-PLZF, which mediate malignant transformation by binding to and dysregulating RARα/RXR and PLZF target genes, respectively. In order to investigate the molecular basis for PLZF-RARα induced leukemogenesis, we used a gain of function model in which PLZF-RARα was ectopically expressed in U937 leukemia cells. After demonstrating in our system that PLZF-RARα is capable of inducing a G1 cell cycle arrest and inhibiting cell growth and myeloid differentiation, we sought to identify genes directly bound and transcriptionally regulated by PLZF-RARα. Chromatin from U937PLZF-RARα expressing cells (+10nM RA) was immunoprecipitated using PLZF antibodies, amplified by ligation-mediated PCR and biological triplicates were hybridized to NimbleGen 2.7kB promoter arrays, which represent 24,275 human promoters. We identified 1797 genes that are directly bound by PLZF-RARα in at least 2 out of 3 arrays, and the majority of these genes (89%) are also bound in the absence of exogenously added RA. Quantitative real time PCR using primary ChIP samples was used to validate ChIP-on-CHIP results and all genes tested to date (n=11) were confirmed as direct targets of PLZF-RARα. Ontological analyses of genes identified by ChIP-on-CHIP revealed enrichment for genes involved in myeloid cell functions including immune, inflammatory and defense responses, in addition to genes involved in apoptosis and signal transduction pathways. Furthermore, genes encoding nuclear proteins were also highly enriched and these included previously identified RARα/RXR target genes (ie. CEBPε, RARβ2, PRAM1, NFE-2), which are likely targeted by the PLZF-RARα oncoprotein, as well as novel PLZF-RARα targets, many of which have roles in blood cell development and have been implicated in leukemia (ie. RUNX1, MLL2, MCL1, PIM1, FANCB). Of these 1797 genes, a significant percentage (22%) are also transcriptionally regulated by PLZF-RARα (>1.5 fold, p<0.05). To identify genes specific to the PLZF-RARα fusion generated in t(11;17) APL, we compared gene expression profiles of 26 PML-RARα and 4 PLZF-RARα expressing APL patient blasts. A comparison of differentially expressed genes in the patient specimens with those both directly bound and regulated by PLZF-RARα in U937 cells, identified a small subset of genes including RUNX1, KLF10, a transcriptional regulator and inhibitor of cell growth, as well as ID1 and ID2, whose expression level has been shown to correlate with myeloid differentiation. Although the expression of these genes was variable in PML-RARα blasts, expression was consistently lower in PLZF-RARα APL blasts (>2 fold, p<0.03). In U937 cells, PLZF-RARα repressed RUNX1, KLF10 and ID1 in the absence of exogenous RA. Intriguingly, RUNX1, KLF10 and ID2 were also identified as direct target genes of PLZF in the KG1a cell line and were transcriptionally regulated by PLZF in U937 cells, suggesting that PLZF and PLZF-RARα may co-regulate a subset of target genes. Given the roles of RUNX1, KLF10, ID1 and ID2 in myeloid differentiation and growth inhibition, these genes may represent PLZF-RARα specific targets that potentially contribute to the pathogenesis of t(11;17) APL.

scholarly journals FLT3 Inhibition Downregulates EZH2 in AML and Promotes Myeloid Differentiation

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 785-785
Author(s):  
Pamela J. Sung ◽  
Simone Sidoli ◽  
Simone S. Riedel ◽  
Katarzyna Kulej ◽  
Hongbo Xie ◽  
...  
Keyword(s):  
Research Funding ◽  
Progenitor Cell ◽  
Target Genes ◽  
Expression Profiles ◽  
Colony Morphology ◽  
Leukemic Cells ◽  
Myeloid Differentiation ◽  
Granulocytic Differentiation ◽  
Induced Differentiation ◽  
Cell State

Abstract Internal tandem duplication mutations in the Fms-like tyrosine kinase 3 (FLT3-ITD) are frequently recurring in AML and confer a poor prognosis. FLT3 inhibitors (FLT3i) such as gilteritinib are efficacious in relapsed AML. Clinical responses to FLT3i include myeloid differentiation of the FLT3-ITD clone in about 50% of patients. How FLT3i induce this response in a subset of patients is unknown. The FLT3i-induced differentiation response seen in clinical trials has not previously been demonstrated in animal models. We modeled FLT3i-induced differentiation in murine Flt3 ITD/ITDDnmt3a -/- AML model (Meyer et al., Cancer Discovery, 2016). Treatment with FLT3i in vitro accelerated differentiation of cKIT+ leukemic splenocytes as assessed by colony morphology in serial re-plating assays. To characterize the differentiation response in vivo, we transplanted CD45.2+ leukemic splenocytes from moribund mice into sub-lethally irradiated healthy congenic CD45.1+ mice. After confirmation of engraftment at 2 weeks post-irradiation, mice were treated with vehicle or gilteritinib for 4 weeks. Animals treated with gilteritinib demonstrated increased neutrophil and decreased stem/progenitor cell populations, recapitulating the clinically observed increase in granulocytic differentiation of the FLT3-ITD clone. We next sought to understand the molecular mechanism of FLT3i-induced differentiation. We used a proteomic-based screen in a human AML cell line treated with FLT3i to identify novel targets of FLT3-ITD that could be potential mediators of the differentiation response. We identified downregulation of Enhancer of Zeste Homolog 2 (EZH2), the catalytic component of the Polycomb Repressive Complex 2 (PRC2). EZH2 and PRC2 were previously shown to be required for leukemic maintenance in mouse models of MLL-AF9 AML. We treated murine Flt3 ITD/ITDDnmt3a -/- cKIT+ leukemic splenocytes with FLT3i or the EZH1/2 inhibitor (EZH1/2i). Both promoted myeloid differentiation to similar degrees as assessed by colony morphology in this model. We hypothesized that FLT3-ITD regulates EZH2 to maintain leukemia cells in a stem/progenitor cell state. We, therefore, characterized the effect of FLT3i on PRC2 in more detail. We confirmed that FLT3i decreases EZH2 protein levels in FLT3-ITD cell lines and primary human AML within 24 hours of treatment as suggested by our proteomic data (Figure 1A-B). We found that the mechanism of EZH2 downregulation is complex with both transcriptional effects and a decrease in EZH2 protein half-life. ChIP-Seq for H3K27me3 demonstrated decreased peaks at the transcription start sites of PRC2 target genes (Figure 1C). RNA-Seq gene expression profiles of FLT3i- and EZH1/2i-treated human AML cells overlapped at 253 differentially expressed genes (Figure 1D). Critically, both FLT3i and EZH1/2i expression profiles enriched in differentiated myeloid cell gene signatures. Overall, we found that EZH2 is a novel, unexpected, and clinically relevant target of FLT3-ITD. Our data suggest that reduced EZH2 activity following FLT3 inhibition promotes myeloid differentiation of FLT3-ITD leukemic cells, providing a mechanistic explanation for the FLT3i-induced differentiation response seen in patients. These data demonstrate that FLT3-ITD has at least two functions in leukemogenesis, the well described activation of signaling pathways, and second, a previously undefined, regulation of PRC2 to maintain a myeloid stem cell state. Our results may lead to improved approaches to therapy for FLT3 mutated AML. Figure 1 Figure 1. Disclosures Bernt: Syndax: Research Funding; Merck: Other: Spouse is an employee of Merck.. Carroll: Incyte Pharmaceuticals: Research Funding; Janssen Pharmaceutical: Consultancy.

scholarly journals E-cigarette use results in suppression of immune and inflammatory-response genes in nasal epithelial cells similar to cigarette smoke

2016 ◽  
Vol 311 (1) ◽  
pp. L135-L144 ◽  
Author(s):  
Elizabeth M. Martin ◽  
Phillip W. Clapp ◽  
Meghan E. Rebuli ◽  
Erica A. Pawlak ◽  
Ellen Glista-Baker ◽  
...  
Keyword(s):  
Cigarette Smoke ◽  
Target Genes ◽  
Smoking Status ◽  
Expression Profiles ◽  
Defense Responses ◽  
Smoking History ◽  
Immune Gene ◽  
Cigarette Smokers ◽  
Human Immunology ◽  
Immune Related Genes

Exposure to cigarette smoke is known to result in impaired host defense responses and immune suppressive effects. However, the effects of new and emerging tobacco products, such as e-cigarettes, on the immune status of the respiratory epithelium are largely unknown. We conducted a clinical study collecting superficial nasal scrape biopsies, nasal lavage, urine, and serum from nonsmokers, cigarette smokers, and e-cigarette users and assessed them for changes in immune gene expression profiles. Smoking status was determined based on a smoking history and a 3- to 4-wk smoking diary and confirmed using serum cotinine and urine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) levels. Total RNA from nasal scrape biopsies was analyzed using the nCounter Human Immunology v2 Expression panel. Smoking cigarettes or vaping e-cigarettes resulted in decreased expression of immune-related genes. All genes with decreased expression in cigarette smokers ( n = 53) were also decreased in e-cigarette smokers. Additionally, vaping e-cigarettes was associated with suppression of a large number of unique genes ( n = 305). Furthermore, the e-cigarette users showed a greater suppression of genes common with those changed in cigarette smokers. This was particularly apparent for suppressed expression of transcription factors, such as EGR1, which was functionally associated with decreased expression of 5 target genes in cigarette smokers and 18 target genes in e-cigarette users. Taken together, these data indicate that vaping e-cigarettes is associated with decreased expression of a large number of immune-related genes, which are consistent with immune suppression at the level of the nasal mucosa.

scholarly journals BCL10 Gain-of-Function Mutations Aberrantly Induce Canonical and Non-Canonical NF-Kb Activation and Resistance to Ibrutinib in ABC-DLBCL

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 2-3
Author(s):  
Min Xia ◽  
Liron David ◽  
Matthew Teater ◽  
Ozlem Onder ◽  
Kojo S. J. Elenitoba-Johnson ◽  
...  
Keyword(s):  
Cell Growth ◽  
Protease Activity ◽  
Research Funding ◽  
Target Genes ◽  
Structural Data ◽  
Gain Of Function ◽  
Spontaneous Polymerization ◽  
Card Domain ◽  
Bcr Signaling ◽  
The Impact

ABC-DLBCLs are among the most aggressive DLBCLs. Genome sequencing studies identified BCL10 gain-of-function mutations in DLBCL mostly within the ABC-DLBCLs. ABC-DLBCLs were recently split into further subtypes according to their genomic characteristics, among which BCL10 somatic mutations were most common in BN2/Cluster 1 cases. These are likely transformed marginal zone lymphomas and were recently suggested to be sensitive to ibrutinib treatment. Pooling publicly available data we noted that most BCL10 mutations are missense or nonsense SNVs affecting its CARD domain, MALT1 binding domain, and the C-terminal S/T rich domain. We generated a large panel of mutant BCL10 constructs including the top hotspot missense mutation R58Q (CARD domain) and nonsense truncation mutations such as E140X. We found that almost all mutants induced aberrantly strong induction of NF-κB activity in lymphoma cells as compared to WT BCL10, indicating that they induce gain of function. BCL10 forms a high order complex with CARD11 and MALT1 (CBM signalosome) downstream of BCR signaling. Normally, CARD11 activation induces polymerization of BCL10 which induces MALT1 activity and downstream NF-κB signaling. To investigate the impact of BCL10 mutants on CBM complex formation we performed fluorescence polarization and filamentation formation assays with purified WT and mutant BCL10 species. Both BCL10R58Q and BCL10E140X manifested faster and even spontaneous polarization compared to BCL10WT. BCL10R58Q formed thicker and more heavily bundled filaments (~20 nm) that provide greater surface area to dock signaling proteins, whereas filaments formed by BCL10E140X had the normal ~10 nm structure. Even though the BCL10E140X deletes the canonical MALT1 binding site, the mutant filament still featured robust MALT1 recruitment. Cryo-EM studies revealed that BCL10R58Q mutant gains new interactions within the filament structure that could explain the observed stabilization and bundling effects. Notably, cryo-EM structure of the BCL10E140X mutant in complex with MALT1 showed that it retains its interaction with MALT1 in the filament form despite its predicted lack of interaction with MALT1 in the monomeric form due to the C-terminal deletion. To gain further functional insight we performed mass spectrometry to identify proteins interacting with WT, BCL10R58Q and BCL10E140X. BCL10R58Q featured gain of many novel protein interactors including NF-κB2 and TAB1 etc. consistent with bundled filament formation enabling more signaling protein recruitment. However, the BCL10E140X interactome was quite different and most notably featured loss of binding to negative regulators of non-canonical NF-κB. NF-κB2 (p100/p52) level was indeed elevated in the presence of this mutant. In addition, both BCL10 mutant classes showed aberrant activation of canonical and non-canonical NF-κB activation (IkBa/p65 and p52) through distinct mechanisms. As a functional readout of BCL10 function, we generated a MALT1 GloSensor reporter DLBCL lines to detect MALT1 protease activity. Indeed, both classes of mutations showed potent induction of MALT1 protease activity (GloSensor), enhanced cleavage of canonical MALT1 target proteins (Western Blot), expression of canonical NF-κB target genes (QPCR) such as IL6 and IL10. In striking contrast to BCL10WT, and consistent with our structural data showing spontaneous polymerization of BCL10 mutants, we found that CARD11 knockdown did not impair MALT1 activation, NF-κB signaling, or cell growth in ABC-DLBCL lines expressing both BCL10 mutants. This CARD11 independence was concerning, since it suggests that BCL10 mutant lymphomas might be resistant to drugs targeting upstream components of the BCR signaling pathway such as ibrutinib. Indeed, expression of BCL10R58Q and BCL10E140X (but not BCL10WT) in various ABC-DLBCL cell lines abrogated the ability of ibrutinib to inhibit MALT1 (GloSensor), NF-κB activity (Reporter), cell growth (Growth inhibition) as well as proliferation. Collectively, we find that BCL10 mutations induce aberrant canonical and non-canonical NF-κB activity through novel and structurally distinct biochemical mechanisms that are at least partially dependent on MALT1. BCL10 mutation should be considered as a biomarker for ibrutinib resistance in ABC-DLBCL, so that alternative targeted therapies can be prioritized for these patients. Disclosures Fontan: Johnson & Johnson: Current Employment. Melnick:Epizyme: Consultancy; Jubilant: Consultancy; Constellation: Consultancy; Janssen: Research Funding; Daiichi Sankyo: Research Funding.

Identification of PML-RARa Target Genes Using Microarray and ChIP-on-Chip Analysis.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2994-2994
Author(s):  
Wenlin Yuan ◽  
Nicole R. Grieselhuber ◽  
Roland D. Green ◽  
Timothy J. Ley
Keyword(s):  
Vector Control ◽  
Cell Lines ◽  
Neutrophil Elastase ◽  
Target Genes ◽  
Experimental System ◽  
Myeloid Cell ◽  
U937 Cells ◽  
Cancer Pathogenesis ◽  
On Chip ◽  
Chip Analysis

Abstract PML-RARa is very toxic to early myeloid cell lines that express neutrophil elastase. As a consequence, myeloid cell lines that stably express PML-RARa must undergo significant adaptations to avoid toxicity (eg. loss of expression of neutrophil elastase (Lane and Ley, MCB, 2005). To identify the direct target genes of PML-RARa, we decided to bypass the issue of adaptation by profiling naive U937 cells that transiently expressed PML-RARa. To define “immediate-early” expression changes caused by PML-RARa, we harvested U937 cells electroporated with expression vectors containing eGFP-PML-RARa (known to possess all the measurable functional properties of PML-RARa) vs. eGFP alone, at the earliest time point when adequate numbers of eGFP positive cells could be collected via high speed cell sorting (6 hours). RNA obtained from these cells was subjected to linear amplification and hybridized with Affymetrix U133 Plus 2.0 arrays. Probesets that demonstrated the largest differences in expression between cells transfected with eGFP vs. eGFP-PML-RARa were PML and RARa themselves (15-fold increase for PML, 9-fold for RARa), providing an important validation for the experimental system. Differentially expressed genes were selected by three filters for further study: (i) ANOVA statistic (P &lt; 0.05); (ii) “present” call and &gt;150 average expression units in at least one sample; and (iii) Reproducible (n=2) &gt;2-fold differences between eGFP-PML-RARa vs. eGFP vector transfected cells. Only 280 probesets (out of 54,613 total) met all of these criteria. Among them, 37 genes were involved in signal transduction, 15 in cell proliferation and differentiation, 7 in apoptosis, 14 in cell cycle, 15 in transport, 27 in protein metabolism and modification, 13 in oncogenesis, and 13 in immunity and defense (Panther classification system). 15/15 genes selected from a variety of categories were further validated with Q-PCR in duplicate using replicate samples, and all differences were statistically significant (p&lt;0.05). Q-PCR validated genes that were up-regulated at 6 hours by eGFP-PML-RARa included CD36 (196% increase over eGFP vector control), LATS2 (204%), Sox4 (395%), COPEB (325%), HoxA5 (294%), IFITM3 (139%), AIF1 (197%), and PLD1 (315%). Q-PCR validated genes that were down-regulated by eGFP-PML-RARa at 6 hours included BTG2 (60% the level of eGFP vector control), KLF7(52%), CDKN1A (p21)(39%), Pim1 (58%), SLC23A2 (36%), HoxA13 (51%), and PDCD1LG1 (42%). Among these genes, Sox4, Pim1, BTG2, and PDCD1LG1 have previously been implicated in cancer pathogenesis by retroviral insertional mutagenesis (http://RTCGD.ncifcrf.gov). To identify promoters that were directly in contact with the eGFP-PML-RARa fusion protein, we performed “ChIP-on-chip” analysis using chromatin immunoprecipitation coupled with a NimbleGen promoter tiling array containing 15 50-mer probes (~100 bp spacing) between positions −1500 and +500 of the transcription initiation sites of a curated subset of 24,275 known human genes (HG17, Build 35). By subtracting non-specific hybridization signals from the eGFP control chip, we identified and validated several promoter regions as direct binding targets of eGFP-PML-RARa, including HoxA13 and CDKN1A (p21). This experimental system provides a new approach for the identification of the direct transcriptional targets of PML-RARa, which may lead to new insights into APL pathogenesis and treatment.

A Comprehensive Genomic Approach Using Gain of Function and Loss of Function Cell Models and ChIP-on-Chip Technology Identifies Novel Promyelocytic Zinc Finger Protein Target Genes.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1407-1407
Author(s):  
Kim L. Rice ◽  
Ari Melnick ◽  
Kenny Ye ◽  
Windy Berkofsky-Fessler ◽  
Jonathan D. Licht
Keyword(s):  
Target Genes ◽  
Rna Binding ◽  
Zinc Finger Protein ◽  
Leukemia Cells ◽  
Loss Of Function ◽  
Function Model ◽  
Gain Of Function ◽  
Hematopoietic Stem ◽  
Endogenous Expression ◽  
On Chip

Abstract The t(11;17)(q23;q21) form of APL involves the production of reciprocal fusion proteins, PLZF-RARα and RARα-PLZF, which mediate malignant transformation by binding to and dysregulating RARα and PLZF target genes. PLZF is expressed in hematopoietic stem cells and is downregulated as cells differentiate. The identification of PLZF target genes including cyclin A2 and MYC is consistent with the hypothesis that PLZF maintains stem cell quiescence by repressing cell cycle driving genes and provides insight into transcriptional pathways disrupted in leukemogenesis. In order to identify additional target genes of PLZF, we constructed a loss of function model in which we suppressed endogenous expression of PLZF using siRNA in KG1a leukemia cells. Our gain of function model consisted of the ectopic expression of PLZF in U937 leukemia cells which do not naturally express PLZF. Expression profiling using GeneChip™ Human Genome U133 Plus 2.0 arrays, which analyze the expression of more than 47,000 transcripts, was performed using both systems. Of the 346 genes identified in the loss of function model, 25% were also regulated by PLZF in the gain of function U937 cell line. Changes in expression of these genes could be direct (through PLZF) or indirect (through secondary effects). In order to determine which genes modulated by changes in PLZF expression are direct transcriptional targets, we immunoprecipitated chromatin using PLZF antibodies in KG1a cells, amplified the products by ligation-mediated PCR and co-hybridized these products with input chromatin to NimbleGen 1.5kB promoter arrays, which represent 24,275 human promoters. Genes bound by PLZF were identified by determining whether consecutively tiled probes were enriched in PLZF-precipitated chromatin as compared to chromatin precipitated with a non-specific antibody. Using a statistical algorithm designed to exclude those probes whose signals of PLZF enrichment might be spuriously identified, we identified 52 genes of the 24,275 on the array as potential PLZF target genes. Strikingly, correlation of these genes with expression analyses revealed that 44% of genes were also significantly regulated by PLZF in the gain of function model and 11% of genes were regulated in the loss of function model. Promoter analyses of a subset of these genes that were identified by ChIP-on-Chip and differentially expressed at least &gt;1.3 fold in PLZF arrays (p&lt;0.05), revealed the presence of a consensus PLZF binding site GTC(C/A)AG in 75% of genes. Analysis of gene ontology for those genes identified by ChIP-on Chip, revealed an enrichment of genes involved in RNA binding and processing as well as genes encoding small G proteins. One gene in particular, RECQL, was directly bound by PLZF in the ChIP-on-Chip assay and transcriptionally regulated by PLZF in both KG1a loss of function and U937 gain of function models. The RECQL protein is a member of the RecQ family of DNA helicases, a class of genes whose mutation is associated with genomic instability tumorigenesis and premature ageing. These data indicate a robust system for the identification of PLZF targets and suggest that PLZF may play a role in genome integrity.

The Predominant microRNAs in β-cell Clusters for Insulin Regulation and Diabetic Control

2020 ◽  
Vol 21 (7) ◽  
pp. 722-734
Author(s):  
Adele Soltani ◽  
Arefeh Jafarian ◽  
Abdolamir Allameh
Keyword(s):  
Mirna Expression ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Diabetic Control ◽  
In Vitro Proliferation ◽  
Cell Functions ◽  
Mirna Expression Profiles ◽  
Multiple Processes ◽  
The Impact

micro (mi)-RNAs are vital regulators of multiple processes including insulin signaling pathways and glucose metabolism. Pancreatic &#946;-cells function is dependent on some miRNAs and their target mRNA, which together form a complex regulative network. Several miRNAs are known to be directly involved in &#946;-cells functions such as insulin expression and secretion. These small RNAs may also play significant roles in the fate of &#946;-cells such as proliferation, differentiation, survival and apoptosis. Among the miRNAs, miR-7, miR-9, miR-375, miR-130 and miR-124 are of particular interest due to being highly expressed in these cells. Under diabetic conditions, although no specific miRNA profile has been noticed, the expression of some miRNAs and their target mRNAs are altered by posttranscriptional mechanisms, exerting diverse signs in the pathobiology of various diabetic complications. The aim of this review article is to discuss miRNAs involved in the process of stem cells differentiation into &#946;-cells, resulting in enhanced &#946;-cell functions with respect to diabetic disorders. This paper will also look into the impact of miRNA expression patterns on in vitro proliferation and differentiation of &#946;-cells. The efficacy of the computational genomics and biochemical analysis to link the changes in miRNA expression profiles of stem cell-derived &#946;-cells to therapeutically relevant outputs will be discussed as well.

scholarly journals Genome-wide transcriptome profiling uncovers differential miRNAs and lncRNAs in ovaries of Hu sheep at different developmental stages

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Samina Shabbir ◽  
Prerona Boruah ◽  
Lingli Xie ◽  
Muhammad Fakhar-e-Alam Kulyar ◽  
Mohsin Nawaz ◽  
...  
Keyword(s):  
Target Genes ◽  
Expression Profiles ◽  
Ovarian Follicle ◽  
Follicular Development ◽  
Differentially Expressed ◽  
Ovary Development ◽  
Estrogen Metabolism ◽  
Genome Wide ◽  
Micro Rnas ◽  
Hu Sheep

AbstractOvary development is an important determinant of the procreative capacity of female animals. Here, we performed genome-wide sequencing of long non-coding RNAs (lncRNAs) and mRNAs on ovaries of 1, 3 and 8 months old Hu sheep to assess their expression profiles and roles in ovarian development. We identified 37,309 lncRNAs, 45,404 messenger RNAs (mRNAs) and 330 novel micro RNAs (miRNAs) from the transcriptomic analysis. Six thousand, seven hundred and sixteen (6716) mRNAs and 1972 lncRNAs were significantly and differentially expressed in ovaries of 1 month and 3 months old Hu sheep (H1 vs H3). These mRNAs and target genes of lncRNAs were primarily enriched in the TGF-β and PI3K-Akt signalling pathways which are closely associated with ovarian follicular development and steroid hormone biosynthesis regulation. We identified MSTRG.162061.1, MSTRG.222844.7, MSTRG.335777.1, MSTRG.334059.16, MSTRG.188947.6 and MSTRG.24344.3 as vital genes in ovary development by regulating CTNNB1, CCNA2, CDK2, CDC20, CDK1 and EGFR expressions. A total of 2903 mRNAs and 636 lncRNAs were differentially expressed in 3 and 8 months old ovaries of Hu sheep (H3 vs H8); and were predominantly enriched in PI3K-Akt, progesterone-mediated oocyte maturation, estrogen metabolism, ovulation from the ovarian follicle and oogenesis pathways. These lncRNAs were also found to regulate FGF7, PRLR, PTK2, AMH and INHBA expressions during follicular development. Our result indicates the identified genes participate in the development of the final stages of follicles and ovary development in Hu sheep.

scholarly journals Dynamic characteristics and functional analysis provide new insights into long non-coding RNA responsive to Verticillium dahliae infection in Gossypium hirsutum

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Guoning Wang ◽  
Xingfen Wang ◽  
Yan Zhang ◽  
Jun Yang ◽  
Zhikun Li ◽  
...  
Keyword(s):  
Disease Resistance ◽  
Verticillium Wilt ◽  
Dynamic Characteristics ◽  
Target Genes ◽  
Acid Metabolism ◽  
Expression Profiles ◽  
Enrichment Analysis ◽  
Alpha Linolenic Acid ◽  
Non Coding Rna ◽  
Long Non Coding Rna

Abstract Background Verticillium wilt is a widespread and destructive disease, which causes serious loss of cotton yield and quality. Long non-coding RNA (lncRNA) is involved in many biological processes, such as plant disease resistance response, through a variety of regulatory mechanisms, but their possible roles in cotton against Verticillium dahliae infection remain largely unclear. Results Here, we measured the transcriptome of resistant G. hirsutum following infection by V. dahliae and 4277 differentially expressed lncRNAs (delncRNAs) were identified. Localization and abundance analysis revealed that delncRNAs were biased distribution on chromosomes. We explored the dynamic characteristics of disease resistance related lncRNAs in chromosome distribution, induced expression profiles, biological function, and these lncRNAs were divided into three categories according to their induced expression profiles. For the delncRNAs, 687 cis-acting pairs and 14,600 trans-acting pairs of lncRNA-mRNA were identified, which indicated that trans-acting was the main way of Verticillium wilt resistance-associated lncRNAs regulating target mRNAs in cotton. Analyzing the regulation pattern of delncRNAs revealed that cis-acting and trans-acting lncRNAs had different ways to influence target genes. Gene Ontology (GO) enrichment analysis revealed that the regulatory function of delncRNAs participated significantly in stimulus response process, kinase activity and plasma membrane components. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis indicated that delncRNAs participated in some important disease resistance pathways, such as plant-pathogen interaction, alpha-linolenic acid metabolism and plant hormone signal transduction. Additionally, 21 delncRNAs and 10 target genes were identified as being involved in alpha-linolenic acid metabolism associated with the biosynthesis of jasmonic acid (JA). Subsequently, we found that GhlncLOX3 might regulate resistance to V. dahliae through modulating the expression of GhLOX3 implicated in JA biosynthesis. Further functional analysis showed that GhlncLOX3-silenced seedlings displayed a reduced resistance to V. dahliae, with down-regulated expression of GhLOX3 and decreased content of JA. Conclusion This study shows the dynamic characteristics of delncRNAs in multiaspect, and suggests that GhlncLOX3-GhLOX3-JA network participates in response to V. dahliae invasion. Our results provide novel insights for genetic improvement of Verticillium wilt resistance in cotton using lncRNAs.

scholarly journals Comprehensive analysis of IgA nephropathy expression profiles: identification of potential biomarkers and therapeutic agents

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Alieh Gholaminejad ◽  
Yousof Gheisari ◽  
Sedigheh Jalali ◽  
Amir Roointan
Keyword(s):  
Iga Nephropathy ◽  
Regulatory Network ◽  
Target Genes ◽  
Expression Profiles ◽  
Gene Signature ◽  
Gene Expression Omnibus ◽  
Integrative Approach ◽  
Potential Factors ◽  
Ngf Signaling ◽  
Underlying Mechanisms

Abstract Background IgA nephropathy (IgAN) is a kidney disease recognized by the presence of IgA antibody depositions in kidneys. The underlying mechanisms of this complicated disease are remained to be explored and still, there is an urgent need for the discovery of noninvasive biomarkers for its diagnosis. In this investigation, an integrative approach was applied to mRNA and miRNA expression profiles in PBMCs to discover a gene signature and novel potential targets/biomarkers in IgAN. Methods Datasets were selected from gene expression omnibus database. After quality control checking, two datasets were analyzed by Limma to identify differentially expressed genes/miRNAs (DEGs and DEmiRs). Following identification of DEmiR-target genes and data integration, intersecting mRNAs were subjected to different bioinformatic analyses. The intersecting mRNAs, DEmiRs, related transcription factors (from TRRUST database), and long-non coding RNAs (from LncTarD database) were used for the construction of a multilayer regulatory network via Cytoscape. Result “GSE25590” (miRNA) and “GSE73953” (mRNA) datasets were analyzed and after integration, 628 intersecting mRNAs were identified. The mRNAs were mainly associated with “Innate immune system”, “Apoptosis”, as well as “NGF signaling” pathways. A multilayer regulatory network was constructed and several hub-DEGs (Tp53, STAT3, Jun, etc.), DEmiRs (miR-124, let-7b, etc.), TFs (NF-kB, etc.), and lncRNAs (HOTAIR, etc.) were introduced as potential factors in the pathogenesis of IgAN. Conclusion Integration of two different expression datasets and construction of a multilayer regulatory network not only provided a deeper insight into the pathogenesis of IgAN, but also introduced several key molecules as potential therapeutic target/non-invasive biomarkers.

Drosophila Gain-of-Function Mutant RTK Torso Triggers Ectopic Dpp and STAT Signaling

Genetics ◽  
2003 ◽  
Vol 164 (1) ◽  
pp. 247-258 ◽  
Author(s):  
Jinghong Li ◽  
Willis X Li
Keyword(s):  
Tyrosine Kinases ◽  
Target Genes ◽  
Tor Signaling ◽  
Gain Of Function ◽  
Essential Requirement ◽  
Downstream Target ◽  
Rtk Signaling ◽  
Genome Wide ◽  
A Genome ◽  
Genomic Regions

Abstract Overactivation of receptor tyrosine kinases (RTKs) has been linked to tumorigenesis. To understand how a hyperactivated RTK functions differently from wild-type RTK, we conducted a genome-wide systematic survey for genes that are required for signaling by a gain-of-function mutant Drosophila RTK Torso (Tor). We screened chromosomal deficiencies for suppression of a gain-of-function mutation tor (torGOF), which led to the identification of 26 genomic regions that, when in half dosage, suppressed the defects caused by torGOF. Testing of candidate genes in these regions revealed many genes known to be involved in Tor signaling (such as those encoding the Ras-MAPK cassette, adaptor and structural molecules of RTK signaling, and downstream target genes of Tor), confirming the specificity of this genetic screen. Importantly, this screen also identified components of the TGFβ (Dpp) and JAK/STAT pathways as being required for TorGOF signaling. Specifically, we found that reducing the dosage of thickveins (tkv), Mothers against dpp (Mad), or STAT92E (aka marelle), respectively, suppressed torGOF phenotypes. Furthermore, we demonstrate that in torGOF embryos, dpp is ectopically expressed and thus may contribute to the patterning defects. These results demonstrate an essential requirement of noncanonical signaling pathways for a persistently activated RTK to cause pathological defects in an organism.

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