scholarly journals Combination of transcriptome sequencing and iTRAQ proteome reveals the molecular mechanisms determining petal shape in herbaceous peony (Paeonia lactiflora Pall.)

2018 ◽  
Vol 38 (6) ◽  
Author(s):  
Yanqing Wu ◽  
Yuhan Tang ◽  
Yu Jiang ◽  
Daqiu Zhao ◽  
Jiali Shang ◽  
...  
Keyword(s):  
Transcriptome Sequencing ◽  
Molecular Mechanisms ◽  
Differentially Expressed ◽  
Paeonia Lactiflora ◽  
Absolute Quantitation ◽  
Herbaceous Peony ◽  
Polymerase Chain ◽  
Petal Shape ◽  
Isobaric Tags ◽  
Rapid Amplification

The molecular mechanisms controlling petal shape in a herbaceous peony, Paeonia lactiflora Pall., a popular high-grade cut flower worldwide, remain unclear. Here, we selected inner and outer petals from P. lactiflora ‘ZiFengyu’ with an anemone type as the study object. Using transcriptome sequencing and isobaric tags for relative and absolute quantitation proteome, 979 differentially expressed genes and 266 differentially expressed proteins were detected within the inner and outer petals. Of these, the present study identified a key gene APETALA2 that regulates flower shape development. In addition, we obtained a 1935 bp full-length cDNA sequence of APETALA2 by rapid amplification of cDNA ends amplification. Through further validation using quantitative real-time polymerase chain reaction and Western blot analysis, APETALA2 showed a markedly higher expression in outer than that in inner petals. Therefore, the present study indicates that the increased expression of APETALA2 contributes to the formation of petals in P. lactiflora.

Isobaric tags for relative and absolute quantitation-based quantitative proteomics analysis provides novel insights into the mechanism of cross-incompatibility between tree peony and herbaceous peony

2019 ◽  
Vol 46 (5) ◽  
pp. 417 ◽  
Author(s):  
Dan He ◽  
Xue-Yuan Lou ◽  
Song-Lin He ◽  
Ya-Kai Lei ◽  
Bo-Va Lv ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Pollen Tube ◽  
Molecular Mechanisms ◽  
Interspecific Hybridisation ◽  
Tree Peony ◽  
Absolute Quantitation ◽  
Herbaceous Peony ◽  
Isobaric Tags ◽  
Shock Proteins

Interspecific hybridisation is the main method for improvement and breeding of tree peony (Paeonia ostii T.Hong & J.X.Zhang), but cross-incompatibility as the major factor restricting the rapid development of interspecific hybridisation. To better understand the molecular mechanisms involved in cross-incompatibility between tree peony (Paeonia ostii cv. Fengdanbai) and herbaceous peony (Paeonia lactiflora Pall. cv. Fenyunu), a quantitative proteomic analysis using isobaric tags for relative and absolute quantitation (iTRAQ) technology was performed on the stigma 24h after pollination. Of the 2900 proteins whose levels were quantitated, 685 proteins were differentially expressed in the stigma after hybrid pollination, in contrast to self-pollination. Functional annotation analysis showed that dysregulated proteins involved in RNA degradation, the Ca signalling pathway, the phosphatidylinositol signalling system and the mitogen-activated protein kinase (MAPK) signalling pathway may have made contributions to cross-incompatibility. The downregulated expression of enolase, DnaK (Heat Shock Proteins, HSP70), GroEL (Heat Shock Proteins, HSP60), calmodulin and glyoxalase I, and the upregulated expression of adenine nucleotide translocator indicated that the energy synthesis required by pollen tube growth, the signal pathway and the metabolic pathway related to the growth polarity of the pollen tube were blocked after hybrid pollination. Eight genes were selected to confirm their expression by quantitative real-time PCR. Compared with the STRING database, a protein–protein interaction network of the chosen proteins was constructed. These results provide fundamental and important information for research into the molecular mechanisms of cross-incompatibility in peony and should facilitate interspecific hybridisation in agricultural practice.

A Comparative Study on the Incidence, Aggravation, and Remission of Lupus Nephritis Based on iTRAQ Technology

2020 ◽  
Vol 23 (7) ◽  
pp. 649-657
Author(s):  
Dong-Jiang Liao ◽  
Xi-Ping Cheng ◽  
Nan Li ◽  
Kang-Li Liang ◽  
Hui Fan ◽  
...  
Keyword(s):  
Lupus Nephritis ◽  
Lupus Erythematosus ◽  
Kidney Tissue ◽  
Quantitative Information ◽  
Enzyme Linked Immunosorbent Assay ◽  
Absolute Quantitation ◽  
Western Blot Method ◽  
Close Relationship ◽  
Polymerase Chain ◽  
Isobaric Tags

Aim and Objective: Lupus nephritis (LN) is one of the major complications of systemic lupus erythematosus (SLE). The specific mechanisms of pathogenesis, aggravation, and remission processes in LN have not been clarified but is of great need in the clinic. Using isobaric tags for relative and absolute quantitation (iTRAQ) technology to screen the functional proteins of LN in mice. Especially under intervention factors of lipopolysaccharide (LPS) and dexamethasone. Methods: Mrl-lps mice were intervened with LPS, dexamethasone, and normal saline (NS) using intraperitoneal injection, and c57 mice intervened with NS as control. The anti-ANA antibody enzyme-linked immunosorbent assay (ELISA) was used to verify disease severity. Kidney tissue is collected and processed for iTRAQ to screen out functional proteins closely related to the onset and development of LN. Western blot method and rt-PCR (real-time Polymerase Chain Reaction) were used for verification. Results: We identified 136 proteins that marked quantitative information. Among them, Hp, Igkv8-27, Itgb2, Got2, and Pcx proteins showed significant abnormal manifestations. Conclusion: Using iTRAQ methods, the functional proteins Hp, Igkv8-27, Itgb2, Got2, and Pcx were screened out for a close relationship with the pathogenesis and development of LN, which is worth further study.

scholarly journals Comparative iTRAQ proteomics revealed proteins associated with lobed fin regeneration in Bichirs

2019 ◽  
Vol 17 (1) ◽  
Author(s):  
Suxiang Lu ◽  
Qian Xiong ◽  
Kang Du ◽  
Xiaoni Gan ◽  
Xuzhen Wang ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Multiple Reaction Monitoring ◽  
Limb Regeneration ◽  
Differentially Expressed ◽  
Receptor Interaction ◽  
Differentially Expressed Proteins ◽  
Absolute Quantitation ◽  
Bioinformatics Analyses ◽  
Fin Regeneration ◽  
Early Stages

Abstract Background Polypterus senegalus can fully regenerate its pectoral lobed fins, including a complex endoskeleton, with remarkable precision. However, despite the enormous potential of this species for use in medical research, its regeneration mechanisms remain largely unknown. Methods To identify the differentially expressed proteins (DEPs) during the early stages of lobed fin regeneration in P. senegalus, we performed a differential proteomic analysis using isobaric tag for relative and absolute quantitation (iTRAQ) approach based quantitative proteome from the pectoral lobed fins at 3 time points. Furthermore, we validated the changes in protein expression with multiple-reaction monitoring (MRM) analysis. Results The experiment yielded a total of 3177 proteins and 15,091 unique peptides including 1006 non-redundant (nr) DEPs. Of these, 592 were upregulated while 349 were downregulated after lobed fin amputation when compared to the original tissue. Bioinformatics analyses showed that the DEPs were mainly associated with Ribosome and RNA transport, metabolic, ECM-receptor interaction, Golgi and endoplasmic reticulum, DNA replication, and Regulation of actin cytoskeleton. Conclusions To our knowledge, this is the first proteomic research to investigate alterations in protein levels and affected pathways in bichirs’ lobe-fin/limb regeneration. In addition, our study demonstrated a highly dynamic regulation during lobed fin regeneration in P. senegalus. These results not only provide a comprehensive dataset on differentially expressed proteins during the early stages of lobe-fin/limb regeneration but also advance our understanding of the molecular mechanisms underlying lobe-fin/limb regeneration.

scholarly journals Identification of differentially expressed proteins in fresh and frozen–thawed boar spermatozoa by iTRAQ-coupled 2D LC–MS/MS

Reproduction ◽  
2014 ◽  
Vol 147 (3) ◽  
pp. 321-330 ◽  
Author(s):  
Xiaoli Chen ◽  
Huabin Zhu ◽  
Chuanhuo Hu ◽  
Haisheng Hao ◽  
Junfang Zhang ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Bioinformatic Analysis ◽  
Absolute Quantification ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Protein Levels ◽  
Boar Semen ◽  
Isobaric Tags ◽  
Boar Spermatozoa

Cryodamage is a major problem in semen cryopreservation, causing changes in the levels of proteins that influence the function and motility of spermatozoa. In this study, protein samples prepared from fresh and frozen–thawed boar spermatozoa were compared using the isobaric tags for relative and absolute quantification (iTRAQ) labeling technique coupled to 2D LC–MS/MS analysis. A total of 41 differentially expressed proteins were identified and quantified, including 35 proteins that were present at higher levels and six proteins that were present at lower levels in frozen–thawed spermatozoa by at least a mean of 1.79-fold (P<0.05). On classifying into ten distinct categories using bioinformatic analysis, most of the 41 differentially expressed proteins were found to be closely relevant to sperm premature capacitation, adhesions, energy supply, and sperm–oocyte binding and fusion. The expression of four of these proteins, SOD1, TPI1, ODF2, and AKAP3, was verified by western blot analysis. We propose that alterations in these identified proteins affect the quality of cryopreserved semen and ultimately lower its fertilizing capacity. This is the first study to compare protein levels in fresh and frozen–thawed spermatozoa using the iTRAQ technology. Our preliminary results provide an overview of the molecular mechanisms of cryodamage in frozen–thawed spermatozoa and theoretical guidance to improve the cryopreservation of boar semen.

scholarly journals Isobaric Tags for Relative and Absolute Quantitation-Based Quantitative Serum Proteomics Analysis in Ischemic Stroke Patients With Hemorrhagic Transformation

2021 ◽  
Vol 15 ◽  
Author(s):  
Zhifeng Qi ◽  
Shuhua Yuan ◽  
Xixi Zhou ◽  
Xunming Ji ◽  
Ke Jian Liu
Keyword(s):  
Ischemic Stroke ◽  
Interaction Analysis ◽  
Hemorrhagic Transformation ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Stroke Patients ◽  
Absolute Quantitation ◽  
Metabolic Pathway Analysis ◽  
Serum Proteomics ◽  
Isobaric Tags

Hemorrhagic transformation (HT), which occurs with or without reperfusion treatments (thrombolysis and/or thrombectomy), deteriorates the outcomes of ischemic stroke patients. It is essential to find clinically reliable biomarkers that can predict HT. In this study, we screened for potential serum biomarkers from an existing blood bank and database with 243 suspected acute ischemic stroke (AIS) patients. A total of 37 patients were enrolled, who were diagnosed as AIS without receiving reperfusion treatment. They were divided into two groups based on whether they were accompanied with HT or not (five HT and 32 non-HT). Serum samples were labeled by isobaric tags for relative and absolute quantitation (iTRAQ) and analyzed by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and compared under NCBInr database. A total of 647 proteins in sera samples were captured, and the levels of 17 proteins (12 upregulated and five downregulated) were significantly different. These differentially expressed proteins were further categorized with Gene Ontology functional classification annotation and Kyoto Encyclopedia of Genes and Genomes metabolic pathway analysis into biological processes. Further protein–protein interaction analysis using String database discovered that, among the differentially expressed proteins, 10 pairs of proteins were found to have crosstalk connections, which may have direct (physical) and indirect (functional) interactions for the development of HT. Our findings suggest that these differentially expressed proteins could serve as potential biomarkers for predicting HT after ischemic stroke.

scholarly journals Quantitative proteomic profiling of Cervicovaginal fluid from pregnant women with term and preterm birth

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Young Eun Kim ◽  
Kwonseong Kim ◽  
Han Bin Oh ◽  
Sung Ki Lee ◽  
Dukjin Kang
Keyword(s):  
Preterm Birth ◽  
Pregnant Women ◽  
Neonatal Morbidity ◽  
Differentially Expressed ◽  
Proteomic Profiling ◽  
Absolute Quantitation ◽  
Term Birth ◽  
Liquid Chromatography Tandem Mass ◽  
Isobaric Tags ◽  
Cervicovaginal Fluid

Abstract Background Preterm birth (PTB) is one of major causes of perinatal mortality and neonatal morbidity, but knowledge of its complex etiology is still limited. Here we present cervicovaginal fluid (CVF) protein profiles of pregnant women who subsequently delivered at spontaneous preterm or term, aiming to identify differentially expressed CVF proteins in PTB and term birth. Methods The CVF proteome of women who sequentially delivered at preterm and term was analyzed using isobaric tags for relative and absolute quantitation (iTRAQ) coupled with two-dimensional nanoflow liquid chromatography-tandem mass spectrometry (2D-nLC-MS/MS). We compared the CVF proteome of PTB (n = 5) and control subjects (term birth, n = 7) using pooled control CVF (term birth, n = 20) as spike-in standard. Results We identified 1294 CVF proteins, of which 605 were newly identified proteins. Of 990 proteins quantified in both PTB and term birth, 52 proteins were significantly up/down-regulated in PTB compared to term birth. The differentially expressed proteins were functionally associated to immune response, endopeptidase inhibitors and structural constituent of cytoskeleton. Finally, we confirm the down-regulation of SERPINB7 (a serine-type protease inhibitor) in PTB compared to control by Western blot. Conclusions Taken together, our study provide quantitative CVF proteome profiles of pregnant women who ultimately delivered at preterm and term. These promising results could help to improve the understanding of PTB etiology and to discover biomarkers for asymptomatic PTB.

scholarly journals Combined Transcriptome and Proteome Analysis of Maize (Zea mays L.) Reveals A Complementary Profile in Response to Phosphate Deficiency

2021 ◽  
Vol 43 (2) ◽  
pp. 1142-1155
Author(s):  
Zhi Nie ◽  
Bowen Luo ◽  
Xiao Zhang ◽  
Ling Wu ◽  
Dan Liu ◽  
...  
Keyword(s):  
Zea Mays ◽  
Zea Mays L ◽  
Proteome Analysis ◽  
Transcript Level ◽  
Utilization Efficiency ◽  
Differentially Expressed ◽  
Phosphate Deficiency ◽  
Absolute Quantitation ◽  
Protein Levels ◽  
Isobaric Tags

A deficiency in the macronutrient phosphate (Pi) brings about various changes in plants at the morphological, physiological and molecular levels. However, the molecular mechanism for regulating Pi homeostasis in response to low-Pi remains poorly understood, particularly in maize (Zea mays L.), which is a staple crop and requires massive amounts of Pi. Therefore, in this study, we performed expression profiling of the shoots and roots of maize seedlings with Pi-tolerant genotype at both the transcriptomic and proteomic levels using RNA sequencing and isobaric tags for relative and absolute quantitation (iTRAQ). We identified 1944 differentially expressed transcripts and 340 differentially expressed proteins under low-Pi conditions. Most of the differentially expressed genes were clustered as regulators, such as transcription factors involved in the Pi signaling pathway at the transcript level. However, the more functional and metabolism-related genes showed expression changes at the protein level. Moreover, under low-Pi conditions, Pi transporters and phosphatases were specifically induced in the roots at both the transcript and protein levels, and increased amounts of mRNA and protein of two purple acid phosphatases (PAPs) and one UDP-sulfoquinovose synthase (SQD) were specifically detected in the roots. The new insights provided by this study will help to improve the P-utilization efficiency of maize.

scholarly journals Transcriptome analysis of two Pogostemon cablin chemotypes reveals genes related to patchouli alcohol biosynthesis

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e12025
Author(s):  
Wuping Yan ◽  
Zhouchen Ye ◽  
Shijia Cao ◽  
Guanglong Yao ◽  
Jing Yu ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Differentially Expressed ◽  
Perennial Herb ◽  
Pcr Analysis ◽  
Biosynthetic Pathways ◽  
Rna Seq ◽  
Patchouli Alcohol ◽  
Pogostemon Cablin ◽  
Polymerase Chain ◽  
Distribution Frequency

Pogostemon cablin, a medicinally and economically important perennial herb, is cultivated around the world due to its medicinal and aromatic properties. Different P. cablin cultivars exhibit different morphological traits and patchouli oil components and contents (especially patchouli alcohol (PA) and pogostone (PO)). According to the signature constituent of the leaf, P. cablin was classified into two different chemotypes, including PA-type and PO-type. To better understand the molecular mechanisms of PA biosynthesis, the transcriptomes of Chinese-cultivated P. cablin cv. PA-type “Nanxiang” (NX) and PO-type “Paixiang” (PX) were analyzed and compared with ribonucleic acid sequencing (RNA-Seq) technology. We obtained a total of 36.83 G clean bases from the two chemotypes, compared them with seven databases and revealed 45,394 annotated unigenes. Thirty-six candidate unigenes participating in the biosynthesis of PA were found in the P. cablin transcriptomes. Overall, 8,390 differentially expressed unigenes were identified between the chemotypes, including 2,467 upregulated and 5,923 downregulated unigenes. Furthermore, six and nine differentially expressed genes (DEGs) were mapped to the terpenoid backbone biosynthetic and sesquiterpenoid and triterpenoid biosynthetic pathways, respectively. One key sesquiterpene synthase gene involved in the sesquiterpenoid and triterpenoid biosynthetic pathways, encoding patchoulol synthase variant 1, was significantly upregulated in NX. Additionally, GC-MS analysis of the two chemotypes in this study showed that the content of PA in NX was significantly higher than that of PX, while the content of PO showed the opposite phenotype. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that the DEG expression tendency was consistent with the transcriptome sequencing results. Overall, 23 AP2/ERF, 13 bHLH, 11 MYB, 11 NAC, three Trihelix, 10 WRKY and three bZIP genes that were differentially expressed may act as regulators of terpenoid biosynthesis. Altogether, 8,314 SSRs were recognized within 6,825 unigenes, with a distribution frequency of 18.32%, among which 1,202 unigenes contained more than one SSR. The transcriptomic characteristics of the two P. cablin chemotypes are comprehensively reported in this study, and these results will contribute to a better understanding of the molecular mechanism of PA biosynthesis. Our transcriptome data also provide a valuable genetic resource for further studies on P. cablin.

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