Transcriptome analysis of transgenic apple fruit overexpressing microRNA172 reveals candidate transcription factors regulating apple fruit development at early stages

Background MicroRNA172 (miR172) has been proven to be critical for fruit growth, since elevated miR172 activity blocks the growth of apple (Malus x domestica Borkh.) fruit. However, it is not clear how overexpression of miR172 affects apple fruit developmental processes. Methods To answer this question, the present study, analyzed global transcriptional changes in miR172-overexpressing (miR172OX) and nongenetically modified wild-type (WT) apple fruit at two developmental stages and in different fruit tissues via RNA-seq. In addition, two cultivars, ‘Hanfu’ and ‘M9’, which have naturally fruit size variation, were included to identify miR172-dependent DEGs. qRT–PCRwas used to verify the reliability of our RNA-seq data. Results Overexpression of miR172 altered the expression levels of many cell proliferation- and cell expansion-related genes. Twenty-four libraries were generated, and 10,338 differentially expressed genes (DEGs) were detected between miR172OX and WT fruit tissues. ‘Hanfu’ and ‘M9’ are two common cultivars that bear fruit of different sizes (250 g and 75 g, respectively). Six libraries were generated, and 3,627 DEGs were detected between ‘Hanfu’ and ‘M9’. After merging the two datasets, 6,888 candidate miR172-specific DEGs were identified. The potential networks associated with fruit size triggered traits were defined among genes belonging to the families of hormone synthesis, signaling pathways, and transcription factors. Our comparative transcriptome analysis provides insights into transcriptome responses to miR172 overexpression in apple fruit and a valuable database for future studies to validate functional genes and elucidate the fruit developmental mechanisms in apple.

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Low Overnight Temperature-Induced Gibberellin Accumulation Increases Locule Number in Tomato

International Journal of Molecular Sciences ◽

10.3390/ijms20123042 ◽

2019 ◽

Vol 20 (12) ◽

pp. 3042

Author(s):

Yanbing Li ◽

Meihua Sun ◽

Hengzuo Xiang ◽

Yudong Liu ◽

Hui Li ◽

...

Keyword(s):

Plant Hormone ◽

Developmental Stages ◽

Fruit Size ◽

Ultra Performance Liquid Chromatography ◽

Rna Seq ◽

Flower Bud ◽

Tomato Plants ◽

Locule Number ◽

Liquid Chromatography Tandem Mass ◽

Flower Bud Differentiation

The number of locules in tomato affects fruit size, shape, and the incidence of malformation. Low temperature increases locule number and the incidences of malformation in tomato plants. In this study, three flower bud developmental stages (pre-flower bud differentiation, sepal and petal primordium formation, and carpel primordium formation) under different night temperatures (10, 15, and 20 °C) were used to analyze the reason behind locule number change using an RNA sequencing (RNA-seq) approach, Quantitative real-time PCR (qRT-PCR), and ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS). The results showed that the “plant hormone signal transduction”, “starch and sucrose metabolism”, and “diterpenoid biosynthesis” categories were remarkably activated during flower bud differentiation. Transcripts of gibberellin (GA)-related genes and endogenous levels of GAs were analyzed, and it was discovered that SlGA2ox genes were significantly downregulated and bioactive GA1 and GA4 accumulated at lower overnight temperature. Exogenous application of bioactive GA1, GA4, and PAC (paclobutrazol) showed that GA1 and GA4 increased the locule number, while PAC decreased the locule number. Taken together, our results suggest that lower overnight temperature reduced the expression of SlGA2ox genes, leading to GA1 and GA4 accumulation, thereby increasing locule number in tomato.

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Endogenous Auxin Content Contributes to Larger Size of Apple Fruit

Frontiers in Plant Science ◽

10.3389/fpls.2020.592540 ◽

2020 ◽

Vol 11 ◽

Author(s):

Haidong Bu ◽

Wenquan Yu ◽

Hui Yuan ◽

Pengtao Yue ◽

Yun Wei ◽

...

Keyword(s):

Regulatory Mechanism ◽

Fruit Size ◽

Apple Fruit ◽

Auxin Signaling ◽

Endogenous Auxin ◽

Rna Seq ◽

Synthetic Genes ◽

Economic Trait ◽

Bud Sport ◽

Auxin Content

Fruit size is an important economic trait that is controlled by multiple genes. However, the regulatory mechanism for fruit size remains poorly understood. A bud sport variety of “Longfeng” (LF) apple (Malus domestica) was identified and named “Grand Longfeng” (GLF). The fruit size of GLF is larger than that of LF, and both varieties are diploid. We found that the cell size in GLF fruit was larger than that of LF. Then, we compared the fruit transcriptomes of the two varieties using RNA-Seq technology. A total of 1166 differentially expressed genes (DEGs) were detected between GLF and LF fruits. The KEGG analysis revealed that the phytohormone pathway was the most enriched, in which most of the DEGs were related to auxin signaling. Moreover, the endogenous auxin levels of GLF fruit were higher than those of LF. The expressions of auxin synthetic genes, including MdTAR1 and MdYUCCA6, were higher in GLF fruit than LF. Collectively, our findings suggest that auxin plays an important role in fruit size development.

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Insights into regulation of C2 and C4 photosynthesis in Amaranthaceae/Chenopodiaceae using RNA-Seq

10.1101/2021.09.14.460237 ◽

2021 ◽

Author(s):

Christian Siadjeu ◽

Maximilian Lauterbach ◽

Gudrun Kadereit

Keyword(s):

Transcription Factors ◽

Transcriptome Analysis ◽

C4 Photosynthesis ◽

Stable State ◽

Comparative Transcriptome ◽

Rna Seq ◽

C4 Species ◽

Genes Encoding ◽

C4 Evolution ◽

Core Genes

Amaranthaceae (incl. Chenopodiaceae) show an immense diversity of C4 syndromes. More than 15 independent origins of C4 photosynthesis, partly in halophytic and/or succulent lineages, and the largest number of C4 species in eudicots signify the importance of this angiosperm lineage in C4 evolution. Here, we conduct RNA-Seq followed by comparative transcriptome analysis of three species from Camphorosmeae representing related clades with different photosynthetic types: Threlkeldiadiffusa (C3), Sedobassiasedoides (C2), and Bassiaprostrata (C4). Results show that B.prostrata belongs to the NADP–ME type and core genes encoding for C4 cycle are significantly up–regulated when compared to Sed.sedoides and T.diffusa, Sedobassiasedoides and B.prostrata share a number of up–regulated C4–related genes, however, two C4 transporters (DIT and TPT) are found significantly up–regulated only in Sed. sedoides. Combined analysis of transcription factors (TFs) of the closely related lineages (Camphorosmeae and Salsoleae) revealed that no C3 specific TFs is higher in C2 species as compared to C4 species, instead the C2 species show their own set of up–regulated TFs. Taken together, our study indicates that the hypothesis of the C2 photosynthesis as a proxy towards C4 photosynthesis is questionable in Sed.sedoides and more in favour of an independent evolutionary stable–state.

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Identification of Transcription Factors Involved in the Regulation of Flowering in Adonis Amurensis Through Combined RNA-seq Transcriptomics and iTRAQ Proteomics

Genes ◽

10.3390/genes10040305 ◽

2019 ◽

Vol 10 (4) ◽

pp. 305 ◽

Cited By ~ 1

Author(s):

Zhou ◽

Sun ◽

Dai ◽

Feng ◽

Zhang ◽

...

Keyword(s):

Transcription Factors ◽

Low Temperatures ◽

Molecular Mechanisms ◽

Developmental Stages ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Global Gene Expression ◽

Early Spring ◽

Rna Seq ◽

Itraq Proteomics

Temperature is one of the most important environmental factors affecting flowering in plants. Adonis amurensis, a perennial herbaceous flower that blooms in early spring in northeast China where the temperature can drop to −15 °C, is an ideal model for studying the molecular mechanisms of flowering at extremely low temperatures. This study first investigated global gene expression profiles at different developmental stages of flowering in A. amurensis by RNA-seq transcriptome and iTRAQ proteomics. Finally, 123 transcription factors (TFs) were detected in both the transcriptome and the proteome. Of these, 66 TFs belonging to 14 families may play a key role in multiple signaling pathways of flowering in A. amurensis. The TFs FAR1, PHD, and B3 may be involved in responses to light and temperature, while SCL, SWI/SNF, ARF, and ERF may be involved in the regulation of hormone balance. SPL may regulate the age pathway. Some members of the TCP, ZFP, MYB, WRKY, and bHLH families may be involved in the transcriptional regulation of flowering genes. The MADS-box TFs are the key regulators of flowering in A. amurensis. Our results provide a direction for understanding the molecular mechanisms of flowering in A. amurensis at low temperatures.

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Transcriptome Analysis of Differentiating Erythroid Progenitors in Refractory Anemia with Ringed Sideroblasts

Blood ◽

10.1182/blood.v116.21.1864.1864 ◽

2010 ◽

Vol 116 (21) ◽

pp. 1864-1864

Author(s):

Per Unneberg ◽

Maryam Nikpour ◽

Liselotte Vesterlund ◽

Monika Jansson ◽

Juha Kere ◽

...

Keyword(s):

Gene Expression ◽

Transcription Factors ◽

Differentially Expressed Genes ◽

Transcriptome Analysis ◽

Expression Profiling ◽

Differentially Expressed ◽

Rna Seq ◽

Gene Model ◽

Time Points ◽

Erythroid Maturation

Abstract Abstract 1864 Refractory anaemia with ring sideroblasts (RARS) represents one subtype of the myelodysplastic syndrome characterized by apoptosis of erythroid precursors and accumulation of aberrant mitochondrial ferritin. Gene expression profiling (GEP) has revealed significant dysregulation of genes involved in iron transport and mitochondrial and erythroid function (Nikpour et al, 2010) but no specific mutations have been identified. Generally, dysregulation increases with forced differentiation indicating alterations of underlying transcription factors. To further explore the molecular mechanisms in RARS, we examined the transcriptional profile associated with early erythroid maturation in NBM and RARS using RNA-Seq. RNA sequencing (RNA-Seq) is a novel method to analyze the full transcriptional activity of a cell or tissue. Expression levels of specific genes, differential splicing, allele-specific expression of transcripts can be accurately determined by RNA-Seq experiments to address many biological-related issues. Bone marrow aspirates were collected from a patient and a control and subjected to CD34+ separation. Cells were cultured for 4 days to allow for erythroid maturation. cDNA libraries were prepared from RNA extracted from these two time points (0 and 4 days), and thereafter sequenced on Life Technology's next generation sequencing platform SOLiD. Approximately 80 million reads were obtained for each library (see Table 1). Two sets of analyses were made; one blinded for position (PU) to allow for an unbiased analysis of data and one (MN) comparing position profiles with raised previous GEP findings. Reads were mapped to the human genome reference sequence (NCBI build version GRCh37) using the bowtie alignment program. To minimize expression level bias through the misclassification of transcripts, only uniquely mapping reads were retained for downstream analysis. Between 14 and 16 million uniquely mapping reads were obtained for each library. Gene models were defined on the genome based on annotations in the ensemble genome database release 58. Reads were associated with a gene if it overlapped any part of a gene, including introns. Furthermore, overlapping genes were removed, ensuring unique read-gene association. 38926 gene models were used in the analysis. Raw gene expression counts for each gene model were obtained as the number of reads that overlapped that gene model. Between 7 and 10 million reads were retained for further analysis. Differential gene expression analysis was done using the bioconductor package DESeq. We compared samples pairwise using a negative binomial statistical model to assess significant differential expression. We corrected for multiple testing using a false discovery rate of 0.1. The number of differentially expressed genes ranged from 10 (comparison Control D4-RARS D4) to 294 (comparison Control CD34+-RARS CD34+). Due to the lack of replicates, variance estimates are uncertain, leading to a lower number of inferred differentially expressed genes. Based on correlation of gene expression and the number of differentially expressed genes, samples could be clustered into two pairs, Control CD34+-RARS CD34+ and Control D4-RARS D4. In order to infer potential functional differences between the samples, we analyzed the function of the differentially expressed genes and performed gene category overrepresentation analysis, using the gene ontology classification system. Interestingly, several non-coding RNAs involved in eg miRNA processing were significantly down-regulated in both RARS positions, compared to NBM. We also identified dramatic dysregulation of two putative zinc finger transcription factors during erythroid differentiation. We conclude that high-quality transcriptome analysis at different time points during erythroid maturation is able to add significant new information to conventional gene expression profiling, which will lead to a more comprehensive understanding of the molecular pathogenesis of RARS. Disclosures: No relevant conflicts of interest to declare.

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Definition of a Small Core Transcriptional Circuit Regulated by AML1-ETO

10.1101/2020.06.14.151159 ◽

2020 ◽

Cited By ~ 1

Author(s):

Kristy R. Stengel ◽

Jacob Ellis ◽

Clare Spielman ◽

Monica Bomber ◽

Scott W. Hiebert

Keyword(s):

Transcription Factors ◽

Gene Networks ◽

Cellular Transformation ◽

Rna Seq ◽

Nascent Transcript ◽

Human Cord Blood ◽

Direct Gene ◽

Transcriptional Changes ◽

Cord Blood Cells ◽

Definition Of

AbstractTranscription factors regulate gene networks controlling normal hematopoiesis and are frequently deregulated in acute myeloid leukemia (AML). Critical to our understanding of the mechanism of cellular transformation by oncogenic transcription factors is the ability to define their direct gene targets. While this seems to be a straight forward task, gene network cascades can change within minutes to hours, making it difficult to distinguish direct from secondary or compensatory transcriptional changes by traditional methodologies. We describe an approach utilizing CRISPR-based genome editing to insert a degron tag into the endogenous AML1-ETO locus of Kasumi-1 cells, as well as overexpression of a degradable AML1-ETO protein in CD34+ human cord blood cells, which is a an AML1-ETO-dependent pre-leukemia model. Upon addition of a small molecule proteolysis targeting chimera (PROTAC), the AML1-ETO protein was rapidly degraded in both systems. Furthermore, by combining rapid degradation with nascent transcript analysis (PRO-seq), RNA-seq and Cut&Run, we have defined the core AML1-ETO regulatory network, which consists of fewer than 100 direct gene targets. The ability of AML1-ETO to regulate this relatively small gene pool is critical for maintaining cells in a self-renewing state, and AML1-ETO degradation set off a cascade of transcriptional events resulting in myeloid differentiation.

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NKL Homeobox Gene VENTX Is Part of a Regulatory Network in Human Conventional Dendritic Cells

International Journal of Molecular Sciences ◽

10.3390/ijms22115902 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5902

Author(s):

Stefan Nagel ◽

Claudia Pommerenke ◽

Corinna Meyer ◽

Hans G. Drexler

Keyword(s):

Dendritic Cells ◽

Transcription Factors ◽

Cell Lines ◽

Expression Profiling ◽

Regulatory Network ◽

Homeobox Gene ◽

Leukemia Cell Line ◽

Specific Gene ◽

Rna Seq ◽

And Function

Recently, we documented a hematopoietic NKL-code mapping physiological expression patterns of NKL homeobox genes in human myelopoiesis including monocytes and their derived dendritic cells (DCs). Here, we enlarge this map to include normal NKL homeobox gene expressions in progenitor-derived DCs. Analysis of public gene expression profiling and RNA-seq datasets containing plasmacytoid and conventional dendritic cells (pDC and cDC) demonstrated HHEX activity in both entities while cDCs additionally expressed VENTX. The consequent aim of our study was to examine regulation and function of VENTX in DCs. We compared profiling data of VENTX-positive cDC and monocytes with VENTX-negative pDC and common myeloid progenitor entities and revealed several differentially expressed genes encoding transcription factors and pathway components, representing potential VENTX regulators. Screening of RNA-seq data for 100 leukemia/lymphoma cell lines identified prominent VENTX expression in an acute myelomonocytic leukemia cell line, MUTZ-3 containing inv(3)(q21q26) and t(12;22)(p13;q11) and representing a model for DC differentiation studies. Furthermore, extended gene analyses indicated that MUTZ-3 is associated with the subtype cDC2. In addition to analysis of public chromatin immune-precipitation data, subsequent knockdown experiments and modulations of signaling pathways in MUTZ-3 and control cell lines confirmed identified candidate transcription factors CEBPB, ETV6, EVI1, GATA2, IRF2, MN1, SPIB, and SPI1 and the CSF-, NOTCH-, and TNFa-pathways as VENTX regulators. Live-cell imaging analyses of MUTZ-3 cells treated for VENTX knockdown excluded impacts on apoptosis or induced alteration of differentiation-associated cell morphology. In contrast, target gene analysis performed by expression profiling of knockdown-treated MUTZ-3 cells revealed VENTX-mediated activation of several cDC-specific genes including CSFR1, EGR2, and MIR10A and inhibition of pDC-specific genes like RUNX2. Taken together, we added NKL homeobox gene activities for progenitor-derived DCs to the NKL-code, showing that VENTX is expressed in cDCs but not in pDCs and forms part of a cDC-specific gene regulatory network operating in DC differentiation and function.

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Transcriptome Analysis of Ginkgo biloba L. Leaves across Late Developmental Stages Based on RNA-Seq and Co-Expression Network

Forests ◽

10.3390/f12030315 ◽

2021 ◽

Vol 12 (3) ◽

pp. 315

Author(s):

Hailin Liu ◽

Xin Han ◽

Jue Ruan ◽

Lian Xu ◽

Bing He

Keyword(s):

Ginkgo Biloba ◽

Leaf Development ◽

Developmental Stages ◽

Leaf Growth ◽

Rna Seq ◽

Final Size ◽

Dioecious Species ◽

Related Factors ◽

New Genes ◽

Gene Modules

The final size of plant leaves is strictly controlled by environmental and genetic factors, which coordinate cell expansion and cell cycle activity in space and time; however, the regulatory mechanisms of leaf growth are still poorly understood. Ginkgo biloba is a dioecious species native to China with medicinally and phylogenetically important characteristics, and its fan-shaped leaves are unique in gymnosperms, while the mechanism of G. biloba leaf development remains unclear. In this study we studied the transcriptome of G. biloba leaves at three developmental stages using high-throughput RNA-seq technology. Approximately 4167 differentially expressed genes (DEGs) were obtained, and a total of 12,137 genes were structure optimized together with 732 new genes identified. More than 50 growth-related factors and gene modules were identified based on DEG and Weighted Gene Co-expression Network Analysis. These results could remarkably expand the existing transcriptome resources of G. biloba, and provide references for subsequent analysis of ginkgo leaf development.

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RNA-Seq Provides New Insights into the Molecular Events Involved in “Ball-Skin versus Bladder Effect” on Fruit Cracking in Litchi

International Journal of Molecular Sciences ◽

10.3390/ijms22010454 ◽

2021 ◽

Vol 22 (1) ◽

pp. 454

Author(s):

Jun Wang ◽

Xiao Fang Wu ◽

Yong Tang ◽

Jian Guo Li ◽

Ming Lei Zhao

Keyword(s):

Transcription Factors ◽

Plant Hormones ◽

Fruit Development ◽

Economic Value ◽

Lipid Synthesis ◽

Expression Level ◽

Agricultural Product ◽

Rna Seq ◽

Fruit Cracking ◽

Molecular Events

Fruit cracking is a disorder of fruit development in response to internal or external cues, which causes a loss in the economic value of fruit. Therefore, exploring the mechanism underlying fruit cracking is of great significance to increase the economic yield of fruit trees. However, the molecular mechanism underlying fruit cracking is still poorly understood. Litchi, as an important tropical and subtropical fruit crop, contributes significantly to the gross agricultural product in Southeast Asia. One important agricultural concern in the litchi industry is that some famous varieties with high economic value such as ‘Nuomici’ are susceptible to fruit cracking. Here, the cracking-susceptible cultivar ‘Nuomici’ and cracking-resistant cultivar ‘Huaizhi’ were selected, and the samples including pericarp and aril during fruit development and cracking were collected for RNA-Seq analysis. Based on weighted gene co-expression network analysis (WGCNA) and the “ball-skin versus bladder effect” theory (fruit cracking occurs upon the aril expanding pressure exceeds the pericarp strength), it was found that seven co-expression modules genes (1733 candidate genes) were closely associated with fruit cracking in ‘Nuomici’. Importantly, we propose that the low expression level of genes related to plant hormones (Auxin, Gibberellins, Ethylene), transcription factors, calcium transport and signaling, and lipid synthesis might decrease the mechanical strength of pericarp in ‘Nuomici’, while high expression level of genes associated with plant hormones (Auxin and abscisic acid), transcription factors, starch/sucrose metabolism, and sugar/water transport might increase the aril expanding pressure, thereby resulting in fruit cracking in ‘Nuomici’. In conclusion, our results provide comprehensive molecular events involved in the “ball-skin versus bladder effect” on fruit cracking in litchi.

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