scholarly journals YBX1 Mediates Alternative Splicing and Maternal mRNA Decay During Pre-Implantation Development

2021 ◽  
Author(s):  
Mingtian Deng ◽  
Baobao Chen ◽  
Zifei Liu ◽  
Yongjie Wan ◽  
Dongxu Li ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Transcriptional Activity ◽  
Mrna Decay ◽  
Molecular Mechanisms ◽  
Rna Stability ◽  
Differentially Expressed ◽  
Rna Seq ◽  
Cell Stage ◽  
Aberrant Expression ◽  
Maternal Mrna

Abstract Background: In mammals, maternal gene products decay and zygotic genome activation (ZGA) during maternal to zygotic transition (MZT) is critical for pre-implantation. Y-box binding protein YBX1 plays vital roles in RNA stabilization and transcriptional regulation, but its roles in pre-implantation development remain to be elucidated. The objective of this study is to investigate the role and the molecular mechanisms of YBX1 during MZT.Methods: RNA-seq datasets in mice, human, bovine, and goat embryos were re-analyzed. YBX1 was knocked down by siRNA microinjection. The 8-cell stage embryos were collected for RNA-seq. The differentially expressed genes and alternative splicing (AS) events were identified using DESeq2 and rMATs, respectively. GO/KEGG/GSEA enrichment analysis was performed using clusterProfiler and enrichplot. Furthermore, 5-EU staining was performed to confirm the effect of YBX1 knockdown on transcriptional activity.Results: The expression of YBX1 was increased during MZT in goat, bovine, human, and mice. By microinjection of siRNA against YBX1, we successfully knocked down YBX1, and the embryo development was impaired in YBX1 knockdown embryos. Using RNA-seq, we identified 1623 up-regulated and 3531 down-regulated genes in the 8-cell stage YBX1 knockdown embryos. Of note, the down-regulated genes were enriched in regulation of RNA/mRNA stability and spliceosome, suggesting that YBX1 might medicate RNA stability and AS. To this end, we identified 3284 differential AS events and 1322 differentially expressed maternal mRNAs at the 8-cell stage YBX1 knockdown embryos. Meanwhile, the splicing factors and mRNA decay related showed aberrant expression. Moreover, the transcriptional activity during ZGA in goat and mice was compromised when YBX1 was knocked down.Conclusion: Our results identify that YBX1 serves an important role in maternal mRNA decay, alternative splicing, and the transcriptional activity required for early embryogenesis, which will broaden the current understanding of YBX1 functions during the stochastic reprogramming events.

scholarly journals Loss of Cnot6l impairs inosine RNA modifications in mouse oocytes

2020 ◽  
Author(s):  
Pavla Brachova ◽  
Nehemiah S. Alvarez ◽  
Lane K. Christenson
Keyword(s):  
Mrna Decay ◽  
Rna Stability ◽  
Adaptor Protein ◽  
Mrna Turnover ◽  
Rna Seq ◽  
Rna Modifications ◽  
Oocyte Competence ◽  
Maternal Mrna ◽  
Maternal Transcripts ◽  
Translation Mechanism

AbstractMammalian oocytes must degrade maternal transcripts through a process called translational mRNA decay, in which maternal mRNA undergoes translational activation, followed by deadenylation and mRNA decay. Once a transcript is translationally activated, it becomes deadenylated by the CCR4-NOT complex. Knockout of Cnot6l, a deadenylase within the CCR4-NOT complex, results in mRNA decay defects during MI entry. Knockout of Btg4, an adaptor protein of the CCR4-NOT complex, results in mRNA decay defects following fertilization. Therefore, mechanisms controlling mRNA turnover have significant impacts on oocyte competence and early embryonic development. Post-transcriptional inosine RNA modifications can impact mRNA stability, possibly through a translation mechanism. Here, we assessed inosine RNA modifications in oocytes from Cnot6l-/- and Btg4-/- mice, which display stabilization of mRNA and over-translation of the stabilized transcripts. If inosine modifications have a role in modulating RNA stability, we hypothesize that in these mutant backgrounds, we would observe changes or a disruption in inosine mRNA modifications. To test this, we used a computational approach to identify inosine RNA modifications in total and polysomal RNA-seq data during meiotic maturation (GV, MI, and MII stages). We observed pronounced depletion of inosine mRNA modifications in oocytes from Cnot6l-/-, but not in Btg4-/- mice. Additionally, analysis of ribosome-associated RNA revealed clearance of inosine modified mRNA. These observations suggest a novel mechanism of mRNA clearance during oocyte maturation, in which inosine-containing transcripts decay in an independent, but parallel mechanism to CCR4-NOT deadenylation.

scholarly journals Changes in Ovary Transcriptome and Alternative splicing at estrus from Xiang pigs with Large and Small Litter Size

2019 ◽  
Author(s):  
Fuping Zhang ◽  
Liangting Tang ◽  
Xueqin Ran ◽  
Ning Mao ◽  
Yiqi Ruan ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Candidate Genes ◽  
Differentially Expressed Genes ◽  
Litter Size ◽  
Molecular Mechanisms ◽  
Reproductive Traits ◽  
Differentially Expressed ◽  
Rna Seq ◽  
Small Litter ◽  
Small Litter Size

AbstractBackground/AimsLitter size is one of the most important reproductive traits in pig breeding, which is affected by multiple genes and the environment. Ovaries are the most important reproductive organs and have a profound impact on the reproduction efficiency. Therefore, genetic differences in the ovaries may contribute to the observed differences in litter size. Although QTLs and candidate genes have been reported to affect the litter size in many pig breeds, however, the findings cannot elucidate the marked differences of the reproductive traits between breeds. The aim of present work is to elucidate the mechanisms of the differences for the reproductive traits and identify candidate genes associated with litter size in Xiang pig breed.MethodsThe changes in ovary transcriptome and alternative splicing were investigated at estrus between Xiang pigs with large and small litter size by RNA-seq technology. The RNA-seq results were confirmed by RT-qPCR method.ResultsWe detected 16,219 - 16,285 expressed genes and 12 types of alternative splicing (AS) events in Xiang pig samples. A total of 762 differentially expressed genes were identified by XL (Xiang pig group with larger litter size) vs XS (Xiang pig group with small litter size) sample comparisons. A total of 34 genes were upregulated and 728 genes were downregulated in XL ovary samples compared with the XS samples. Alternative splicing (AS) rates in XL samples were slightly lower than that observed in XS samples. Most of differentially expressed genes were differentially regulated on AS level. Eleven candidate genes were potentially identified to be related to Xiang pig fecundity and litter size, which may be closely related to the gonad development, oocyte maturation or embryo quality.ConclusionThe significant changes in the expression of the protein-coding genes and the level of alternative splicing in estrus ovarian transcriptome between XL and XS groups probably are the molecular mechanisms of phenotypic variation in litter size.

scholarly journals Loss of Cnot6l Impairs Inosine RNA Modifications in Mouse Oocytes

2021 ◽  
Vol 22 (3) ◽  
pp. 1191
Author(s):  
Pavla Brachova ◽  
Nehemiah S. Alvarez ◽  
Lane K. Christenson
Keyword(s):  
Mrna Decay ◽  
Rna Stability ◽  
Adaptor Protein ◽  
Rna Seq ◽  
Rna Modifications ◽  
Oocyte Competence ◽  
Maternal Mrna ◽  
Maternal Transcripts ◽  
Translation Mechanism ◽  
Gene 4

Mammalian oocytes must degrade maternal transcripts through a process called translational mRNA decay, in which maternal mRNA undergoes translational activation, followed by deadenylation and mRNA decay. Once a transcript is translationally activated, it becomes deadenylated by the CCR4-NOT complex. Knockout of CCR4-NOT Transcription Complex Subunit 6 Like (Cnot6l), a deadenylase within the CCR4-NOT complex, results in mRNA decay defects during metaphase I (MI) entry. Knockout of B-cell translocation gene-4 (Btg4), an adaptor protein of the CCR4-NOT complex, results in mRNA decay defects following fertilization. Therefore, mechanisms controlling mRNA turnover have significant impacts on oocyte competence and early embryonic development. Post-transcriptional inosine RNA modifications can impact mRNA stability, possibly through a translation mechanism. Here, we assessed inosine RNA modifications in oocytes, eggs, and embryos from Cnot6l-/- and Btg4-/- mice, which display stabilization of mRNA and over-translation of the stabilized transcripts. If inosine modifications have a role in modulating RNA stability, we hypothesize that in these mutant backgrounds, we would observe changes or a disruption in inosine mRNA modifications. To test this, we used a computational approach to identify inosine RNA modifications in total and polysomal RNA-seq data during meiotic maturation (GV, MI, and MII stages). We observed pronounced depletion of inosine mRNA modifications in samples from Cnot6l-/-, but not in Btg4-/- mice. Additionally, analysis of ribosome-associated RNA revealed clearance of inosine modified mRNA. These observations suggest a novel mechanism of mRNA clearance during oocyte maturation, in which inosine-containing transcripts decay in an independent, but parallel mechanism to CCR4-NOT deadenylation.

scholarly journals Comparative Transcriptome Analysis Reveals Sex-Based Differences during the Development of the Adult Parasitic Wasp Cotesia vestalis (Hymenoptera: Braconidae)

Genes ◽  
2021 ◽  
Vol 12 (6) ◽  
pp. 896
Author(s):  
Yuenan Zhou ◽  
Pei Yang ◽  
Shuang Xie ◽  
Min Shi ◽  
Jianhua Huang ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Adult Development ◽  
Sexual Development ◽  
Molecular Mechanisms ◽  
Parasitic Wasp ◽  
Differentially Expressed Gene ◽  
Regulation Mechanism ◽  
Rna Seq ◽  
Male And Female ◽  
Cotesia Vestalis

The endoparasitic wasp Cotesia vestalis is an important biological agent for controlling the population of Plutella xylostella, a major pest of cruciferous crops worldwide. Though the genome of C. vestalis has recently been reported, molecular mechanisms associated with sexual development have not been comprehensively studied. Here, we combined PacBio Iso-Seq and Illumina RNA-Seq to perform genome-wide profiling of pharate adult and adult development of male and female C. vestalis. Taking advantage of Iso-Seq full-length reads, we identified 14,466 novel transcripts as well as 8770 lncRNAs, with many lncRNAs showing a sex- and stage-specific expression pattern. The differentially expressed gene (DEG) analyses showed 2125 stage-specific and 326 sex-specific expressed genes. We also found that 4819 genes showed 11,856 alternative splicing events through combining the Iso-Seq and RNA-Seq data. The results of comparative analyses showed that most genes were alternatively spliced across developmental stages, and alternative splicing (AS) events were more prevalent in females than in males. Furthermore, we identified six sex-determining genes in this parasitic wasp and verified their sex-specific alternative splicing profiles. Specifically, the characterization of feminizer and doublesex splicing between male and female implies a conserved regulation mechanism of sexual development in parasitic wasps.

scholarly journals Maternal factor PABPN1L is essential for maternal mRNA degradation during maternal-to-zygotic transition

2020 ◽  
Author(s):  
Ying Wang ◽  
Tianhao Feng ◽  
Xiaodan Shi ◽  
Siyu Liu ◽  
Zerui Wang ◽  
...  
Keyword(s):  
Mouse Model ◽  
Large Scale ◽  
Mrna Degradation ◽  
Female Infertility ◽  
Control Group ◽  
Rna Seq ◽  
Cell Stage ◽  
Maternal Mrna ◽  
Maternal To Zygotic Transition ◽  
Group A

AbstractInfertility affects 10% - 15% of families worldwide. However, the pathogenesis of female infertility caused by abnormal early embryonic development is not clear. We constructed a mouse model (Pabpn1l -/-) simulating the splicing abnormality of human PABPN1L and found that the female was sterile and the male was fertile. The Pabpn1l -/- oocytes can be produced, ovulated and fertilized normally, but cannot develop beyond the 2-cell stage. Using RNA-Seq, we found a large-scale upregulation of RNA in Pabpn1l -/- MII oocytes. Of the 2401 transcripts upregulated in Pabpn1l-/- MII oocytes, 1523 transcripts (63.4%) were also upregulated in Btg4 -/- MII oocytes, while only 53 transcripts (2.2%) were upregulated in Ythdf2 -/- MII oocytes. We documented that transcripts in zygotes derived from Pabpn1l -/- oocytes have a longer poly(A) tail than the control group, a phenomenon similar to that in Btg4-/- mice. Surprisingly, the poly(A) tail of these mRNAs was significantly shorter in the Pabpn1l -/- MII oocytes than in the Pabpn1l +/+. These results suggest that PABPN1L is involved in BTG4-mediated maternal mRNA degradation, and may antagonize poly(A) tail shortening in oocytes independently of its involvement in maternal mRNA degradation. Thus, PABPN1L variants could be a genetic marker of female infertility.

scholarly journals Alternative splicing of leptin receptor overlapping transcript in osteosarcoma

2020 ◽  
Vol 245 (16) ◽  
pp. 1437-1443
Author(s):  
Emel Rothzerg ◽  
Xuan D Ho ◽  
Jiake Xu ◽  
David Wood ◽  
Aare Märtson ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Molecular Mechanisms ◽  
Leptin Receptor ◽  
Osteogenic Sarcoma ◽  
Differentially Expressed ◽  
Primary Malignancy ◽  
Transcript Variants ◽  
Potential Biomarker ◽  
Whole Transcriptome Analysis ◽  
Tumor Biopsies

Alternative splicing of RNA is an essential mechanism that increases proteomic diversity in eukaryotic cells. Aberrant alternative splicing is often associated with various human diseases, including cancer. We conducted whole-transcriptome analysis of 18 osteosarcoma bone samples (paired normal—tumor biopsies). Using RNA-seq, we identified statistically significant (FDR <0.05) 26 differentially expressed transcript variants of leptin receptor overlapping transcript ( LEPROT) gene. Some of the transcripts were overexpressed in normal cells, whereas others were overexpressed in tumor cells. The function of LEPROT is not completely understood. Herein, we highlight a possible association between OS and aberrant alternative splicing events and its interaction with the expression of LEPROT. We also discuss the role of LEPROT in regulating growth hormone and its receptor, and the relationship with initiation and progression of OS. This research study may help to understand the association of alternative splicing mechanism in OS and in tumorigenesis more generally. Further, LEPROT gene can also be considered as a potential biomarker of osteosarcoma. Impact statement Osteosarcoma (OS, also known as osteogenic sarcoma) is the most common primary malignancy of bone in children and adolescents. The molecular mechanisms of OS are extremely complicated and its molecular mediators remain to be elucidated. We sequenced total RNA from 18 OS bone samples (paired normal—tumor biopsies). We found statistically significant (FDR <0.05) 26 differentially expressed transcript variants of LEPROT gene with different expressions in normal and tumor samples. These findings contribute to the understanding of molecular mechanisms of OS development and provide encouragement to pursue further research.

scholarly journals Integration of mRNA and miRNA Analysis Reveals the Molecular Mechanism Underlying Salt and Alkali Stress Tolerance in Tobacco

2019 ◽  
Vol 20 (10) ◽  
pp. 2391 ◽  
Author(s):  
Jiayang Xu ◽  
Qiansi Chen ◽  
Pingping Liu ◽  
Wei Jia ◽  
Zheng Chen ◽  
...  
Keyword(s):  
Salinity Stress ◽  
Regulatory Networks ◽  
Molecular Mechanisms ◽  
Crop Yields ◽  
Antioxidant Activities ◽  
Expression Patterns ◽  
Differentially Expressed ◽  
Alkali Stress ◽  
Rna Seq ◽  
Alkali Tolerance

Salinity is one of the most severe forms of abiotic stress and affects crop yields worldwide. Plants respond to salinity stress via a sophisticated mechanism at the physiological, transcriptional and metabolic levels. However, the molecular regulatory networks involved in salt and alkali tolerance have not yet been elucidated. We developed an RNA-seq technique to perform mRNA and small RNA (sRNA) sequencing of plants under salt (NaCl) and alkali (NaHCO3) stress in tobacco. Overall, 8064 differentially expressed genes (DEGs) and 33 differentially expressed microRNAs (DE miRNAs) were identified in response to salt and alkali stress. A total of 1578 overlapping DEGs, which exhibit the same expression patterns and are involved in ion channel, aquaporin (AQP) and antioxidant activities, were identified. Furthermore, genes involved in several biological processes, such as “photosynthesis” and “starch and sucrose metabolism,” were specifically enriched under NaHCO3 treatment. We also identified 15 and 22 miRNAs that were differentially expressed in response to NaCl and NaHCO3, respectively. Analysis of inverse correlations between miRNAs and target mRNAs revealed 26 mRNA-miRNA interactions under NaCl treatment and 139 mRNA-miRNA interactions under NaHCO3 treatment. This study provides new insights into the molecular mechanisms underlying the response of tobacco to salinity stress.

RNA-seq analysis of 2 closely related leukemia clones that differ in their self-renewal capacity

Blood ◽  
2011 ◽  
Vol 117 (2) ◽  
pp. e27-e38 ◽  
Author(s):  
Brian T. Wilhelm ◽  
Mathieu Briau ◽  
Pamela Austin ◽  
Amélie Faubert ◽  
Geneviève Boucher ◽  
...  
Keyword(s):  
Stem Cell ◽  
Molecular Mechanisms ◽  
Differentially Expressed ◽  
Leukemic Cells ◽  
Rna Seq ◽  
Self Renewal ◽  
Leukemia Stem Cell ◽  
Cellular Behavior ◽  
Transcriptome Variation ◽  
G Protein Coupled

Abstract The molecular mechanisms regulating self-renewal of leukemia stem cells remain poorly understood. Here we report the generation of 2 closely related leukemias created through the retroviral overexpression of Meis1 and Hoxa9. Despite their apparent common origin, these clonal leukemias exhibit enormous differences in stem cell frequency (from 1 in 1.4, FLA2; to 1 in 347, FLB1), suggesting that one of these leukemias undergoes nearly unlimited self-renewal divisions. Using next-generation RNA-sequencing, we characterized the transcriptomes of these phenotypically similar, but biologically distinct, leukemias, identifying hundreds of differentially expressed genes and a large number of structural differences (eg, alternative splicing and promoter usage). Focusing on ligand-receptor pairs, we observed high expression levels of Sdf1-Cxcr4; Jagged2-Notch2/1; Osm-Gp130; Scf-cKit; and Bmp15-Tgfb1/2. Interestingly, the integrin beta 2-like gene (Itgb2l) is both highly expressed and differentially expressed between our 2 leukemias (∼ 14-fold higher in FLA2 than FLB1). In addition, gene ontology analysis indicated G-protein-coupled receptor had a much higher proportion of differential expression (22%) compared with other classes (∼ 5%), suggesting a potential role regulating subtle changes in cellular behavior. These results provide the first comprehensive transcriptome analysis of a leukemia stem cell and document an unexpected level of transcriptome variation between phenotypically similar leukemic cells.

scholarly journals RNA-Seq Profiling of Circular RNAs During Development of Hindgut in Rat Embryos With Ethylenethiourea-Induced Anorectal Malformations

2021 ◽  
Vol 12 ◽  
Author(s):  
Si Ying Li ◽  
Chen Yi Wang ◽  
Yun Xia Xiao ◽  
Xiao Bing Tang ◽  
Zheng Wei Yuan ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Transforming Growth Factor ◽  
Quantitative Polymerase Chain Reaction ◽  
Anorectal Malformations ◽  
Normal Group ◽  
Differentially Expressed ◽  
Mitogen Activated Protein Kinase ◽  
Circular Rnas ◽  
Rna Seq ◽  
Pregnant Rats

Anorectal malformations (ARMs) are among the most common congenital terminal digestive tract malformations. Circular RNAs (circRNAs), a novel type of endogenous non-coding RNAs, play roles in the development of the digestive system; however, their contributions to the pathogenesis of ARMs are not well-established. In this study, we explored the mechanism underlying ethylenethiourea (ETU)-induced ARMs by profiling circRNA expression via RNA-seq and constructing a regulatory circRNA-miRNA-mRNA network. Nine pregnant rats were gavage-fed a single dose of 125 mg/kg 1% ETU (ARM group) on gestational day 10 (GD10), and another 9 pregnant rats received a similar dose of saline (normal group) as a control. Embryos were obtained by cesarean section on the key time-points of anorectal development (GD14, GD15, and GD16). Hindgut samples isolated from the fetuses were evaluated by high-throughput sequencing and differentially expressed circRNAs were validated by reverse transcription-quantitative polymerase chain reaction, agarose gel electrophoresis, and Sanger cloning and sequencing. A total of 18295 circRNAs were identified in the normal and ARM groups. Based on the 425 differentially expressed circRNAs (|Fc| &gt; 2, p &lt; 0.05), circRNA-miRNA and miRNA-mRNA pairs were predicted using miREAP, miRanda, and TargetScan. A total of 55 circRNAs (14 up- and 41 downregulated in the ARM group compared to the normal group) were predicted to bind to 195 miRNAs and 947 mRNAs. Competing endogenous RNA networks and a Kyoto Encyclopedia of Genes and Genomes analysis revealed that novel_circ_001042 had the greatest connectivity and was closely related to ARM-associated signaling pathways, such as the Wingless Type MMTV integration site family, mitogen-activated protein kinase, and transforming growth factor-β pathways. These results provide original insight into the roles of circRNAs in ARMs and provide a valuable resource for further analyses of molecular mechanisms and signaling networks.

scholarly journals Molecular Mechanisms Underlying Sugarcane Response to Aluminum Stress by RNA-Seq

2020 ◽  
Author(s):  
Thiago Mateus Rosa-Santos ◽  
Renan Gonçalves da Silva ◽  
Poornasree Kumar ◽  
Pratibha Kottapalli ◽  
Chiquito Crasto ◽  
...  
Keyword(s):  
Differentially Expressed Genes ◽  
Molecular Mechanisms ◽  
Aluminum Tolerance ◽  
Differentially Expressed ◽  
Rna Seq ◽  
Aluminum Stress ◽  
Physiological Processes ◽  
Aluminum Ions ◽  
Detoxification Mechanism ◽  
Genomic Studies

Sugarcane is an important sugar-source crop. As any other plant, it can be exposed to several abiotic stress conditions. Though some metals contribute to critical physiological processes in plants, the presence of aluminum ions (Al3+) can be very toxic. In order to develop plants that flourish in acidic soils, it is critical to gain insights into the molecular mechanisms of sugarcane response to aluminum stress. To determine the genes involved in sugarcane response to aluminum stress we generated 372 million paired-end RNA sequencing reads, from roots of CTC-2 and RB855453 two contrasting cultivars. Data normalization resulted in 162,161 contigs and 97,335 trinity genes. After the read cutoff, the differentially expressed genes were 4,858 in CTC-2 and 1,307 in the RB855453, Treatment Vs Control, respectively. The differentially expressed genes were annotated into 34 functional categories. The majority of the genes were upregulated in the CTC-2 (tolerant cultivar) and down regulated in RB855453 (sensitive cultivar). Here, we present the first root-transcriptome of sugarcane under aluminum stress. The results and conclusions of this study provide a valuable resource for future genetic and genomic studies in sugarcane. This transcriptome analysis points out that sugarcane tolerance to aluminum may be explained by an efficient detoxification mechanism combined with the lateral root formation and activation of redox enzymes. Following our results, we present here, a hypothetical model for the aluminum tolerance in CTC-2 cultivar.

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