iTRAQ-Based Proteomics Reveals Novel Members Involved in Pathogen Challenge in Sea Cucumber Apostichopus japonicus

Zhang, Pengjuan; Li, Chenghua; Zhang, Peng; Jin, Chunhua; Pan, Daodong; Bao, Yongbo

doi:10.1371/journal.pone.0100492

Cited by 46 publications

(33 citation statements)

References 34 publications

(38 reference statements)

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“…As shown in Figure 2, a 5.53-and 7.64-fold increase was detected at 48 hr and 96 hr in bacteria-infected samples, respectively, compared with the control group (Figure 2A), and a 3.06-fold increase was detected at 6 hr after exposure to 1 mgÁml 21 LPS ( Figure 2B). However, the AjBHMT protein level initially increased by 4.01-fold at 24 hr postinfection, but gradually decreased to control levels at 48 hr and continuously declined at 96 hr in V. splendidus-challenged sea cucumbers from our iTRAQ qualification (Zhang et al 2014) ( Figure 2C). These findings indicated that AjBHMT should be tightly regulated at different stages of infection.…”

Section: Smart Analysis Indicated Thatmentioning

confidence: 85%

The Roles of Two miRNAs in Regulating the Immune Response of Sea Cucumber

Zhang

et al. 2015

Genetics

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MicroRNAs (miRNAs) have emerged as key regulators in many pathological processes by suppressing the transcriptional and post-transcriptional expression of target genes. MiR-2008 was previously found to be significantly up-regulated in diseased sea cucumber Apostichopus japonicus by high-through sequencing, whereas the reads of miR-137, a well-documented tumor repressor, displayed no significant change. In the present study, we found that miR-137 expression was slightly attenuated and miR-2008 was significantly enhanced after Vibrio splendidus infection or Lipopolysaccharides application. Further target screening and dual-luciferase reporter assay revealed that the two important miRNAs shared a common target gene of betaine-homocysteine S-methyltransferase (AjBHMT), which exhibited noncorrelated messenger RNA and protein expression patterns after bacterial challenge. In order to fully understand their regulatory mechanisms, we conducted the functional experiments in vitro and in vivo. The overexpression of miR-137 in sea cucumber or primary coelomocytes significantly decreased, whereas the inhibition of miR-137 increased the mRNA and protein expression levels of AjBHMT. In contrast, miR-2008 overexpression and inhibition showed no effect on AjBHMT mRNA levels, but the concentration of AjBHMT protein displayed significant changes both in vitro and in vivo. Consistently, the homocysteine (Hcy) contents were also accordingly altered in the aberrant expression analysis of both miRNAs, consistent with the results of the AjBHMT silencing assay in vitro and in vivo. More importantly, small interfering RNA mediated AjBHMT knockdown and Hcy exposure analyses both significantly increased reactive oxygen species (ROS) production and decreased the number of surviving invasive pathogen in sea cucumber coelomocytes. Taken together, these findings confirmed the differential roles of sea cucumber miR-137 and miR-2008 in regulating the common target AjBHMT to promote ROS production and the clearance of pathogenic microorganisms through Hcy accumulation.KEYWORDS Apostichopus japonicus; miR-137; miR-2008; betaine-homocysteine S-methyltransferase; homocysteine; ROS, genetics of immunity M ICRORNAS (miRNAs) are a class of endogenous small noncoding RNAs of $22 nucleotides, which typically function as repressors of target genes, resulting in direct messenger RNA (mRNA) degradation or translation inhibition (Bartel 2004). These RNAs play crucial roles in many cellular processes, such as proliferation, differentiation, apoptosis, and survival (Lai 2002;He and Hannon 2004;Miska 2005). The expression of up to 60% of all protein-coding genes is estimated to be under the control of miRNAs (Friedman 2009), and combinatorial regulation has been identified as a prominent feature of miRNA regulation. Thus, a given miR-NA might have multiple different mRNA targets, and a given mRNA might similarly be targeted by multiple miRNAs (Krek et al. 2005;Rajewsky 2006). Consistent with the crucial roles of miRNA in the normal functioning of euka...

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Section: Smart Analysis Indicated Thatmentioning

confidence: 85%

The Roles of Two miRNAs in Regulating the Immune Response of Sea Cucumber

Zhang

et al. 2015

Genetics

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“…Peptide fractionation and mass-spectrometry analysis were routinely carried out as described by Zhang et al [34], and the enzymatically hydrolyzed protein solution was re-suspended with 110 µL of eluent A (acetonitrile-H 2 O-formic acid, 2:98:0.1, v/v/v). Peptides were separated using an Agilent 1200 HPLC (Wilmington, DE, USA) with a guard column and a separation column, and UV detection wavelengths were 210 nm and 280 nm.…”

Section: Peptide Fractionation and Mass-spectrometry Analysismentioning

confidence: 99%

“…The MS/MS spectra were processed using Protein Pilot Software v.5.0 (AB Sciex, Framingham, MA, USA) as described by Zhang et al [34]. Laconically, only peptides with the 95% confidence interval were counted as the identified protein.…”

Section: Protein Identification and Quantificationmentioning

confidence: 99%

Cuminal Inhibits Trichothecium roseum Growth by Triggering Cell Starvation: Transcriptome and Proteome Analysis

Zhang

et al. 2020

Microorganisms

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Trichothecium roseum is a harmful postharvest fungus causing serious damage, together with the secretion of insidious mycotoxins, on apples, melons, and other important fruits. Cuminal, a predominant component of Cuminum cyminum essential oil has proven to successfully inhibit the growth of T. roseum in vitro and in vivo. Electron microscopic observations revealed cuminal exposure impaired the fungal morphology and ultrastructure, particularly the plasmalemma. Transcriptome and proteome analysis was used to investigate the responses of T. roseum to exposure of cuminal. In total, 2825 differentially expressed transcripts (1516 up and 1309 down) and 225 differentially expressed proteins (90 up and 135 down) were determined. Overall, notable parts of these differentially expressed genes functionally belong to subcellular localities of the membrane system and cytosol, along with ribosomes, mitochondria and peroxisomes. According to the localization analysis and the biological annotation of these genes, carbohydrate and lipids metabolism, redox homeostasis, and asexual reproduction were among the most enriched gene ontology (GO) terms. Biological pathway enrichment analysis showed that lipids and amino acid degradation, ATP-binding cassette transporters, membrane reconstitution, mRNA surveillance pathway and peroxisome were elevated, whereas secondary metabolite biosynthesis, cell cycle, and glycolysis/gluconeogenesis were down regulated. Further integrated omics analysis showed that cuminal exposure first impaired the polarity of the cytoplasmic membrane and then triggered the reconstitution and dysfunction of fungal plasmalemma, resulting in handicapped nutrient procurement of the cells. Consequently, fungal cells showed starvation stress with limited carbohydrate metabolism, resulting a metabolic shift to catabolism of the cell's own components in response to the stress. Additionally, these predicaments brought about oxidative stress, which, in collaboration with the starvation, damaged certain critical organelles such as mitochondria. Such degeneration, accompanied by energy deficiency, suppressed the biosynthesis of essential proteins and inhibited fungal growth.fungicides to be used for postharvest treatment, reduced efficacy due to the emergence of pathogen resistance to these chemicals, and the growing public concerns over chemical residues in food and the environment have necessitated the seeking of alternative solutions to control the fungal spoilage and to guarantee food safety and consumer health [8].Essential oils (EOs), generally recognized as safe by the US Food and Drug Administration [9], are natural substances with established antimicrobial activities [10,11] and are a candidate instrument for controlling postharvest disease [12,13]. Nonetheless, this candidacy is compromised by the expensiveness of these natural substances due to their generally low yields. However, exploring their mode of actions is valuable to the development of novel alternatives.The multicomponent EOs attribute their mode of act...

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“…Recently, proteomic studies have become very popular, especially for non‐model organisms and those lacking reference genomes (Meng, Hou, Zhao, Huang, Huang, Xia & Wang ; Zhang, Li, Zhang, Jin, Pan & Bao ). In most of the studies, proteome analysis was conducted on the basis of transcriptome analysis as it was thought that transcriptome‐deduced proteins could cover almost all the proteins found using the isobaric tags for relative and absolute quantitation (iTRAQ) method.…”

Section: Introductionmentioning

confidence: 99%

iTRAQ-based identification of differentially expressed proteins related to growth in the swimming crab, Portunus trituberculatus

Ren

Gao

et al. 2016

Aquac Res

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The swimming crab, Portunus trituberculatus, is an important farmed species in China, and it has been studied extensively, which requires more information on its genetic background. To date, information on the proteomics of P. trituberculatus is scarce. In this study, we used isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomics to investigate growth regulation in P. trituberculatus. Total proteins were isolated from five tissues (eyestalk, gill, heart, hepatopancreas and muscle). Equal quantities of protein from each tissue were pooled at the proteome level using the iTRAQ method. A total of 961 proteins were identified using liquid chromatography coupled with tandem mass spectrometry and de novo sequencing data. Using a 1.2-fold change in expression as a physiologically significant benchmark, 30 differentially expressed proteins (DEPs) were found to undergo differential expression related to the growth of P. trituberculatus, and most of the growth-related proteins, including those involved in metabolism, immune responses, DNA duplication and protein synthesis, were upregulated, indicating that conservation of energy is an important strategy to cope with growth. There was a high consistency between the expression levels determined using iTRAQ and mRNA, highlighting the high reproducibility of our proteomic approach and its great value in elucidating the molecular mechanisms of P. trituberculatus.

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iTRAQ-Based Proteomics Reveals Novel Members Involved in Pathogen Challenge in Sea Cucumber Apostichopus japonicus

Cited by 46 publications

References 34 publications

The Roles of Two miRNAs in Regulating the Immune Response of Sea Cucumber

The Roles of Two miRNAs in Regulating the Immune Response of Sea Cucumber

Cuminal Inhibits Trichothecium roseum Growth by Triggering Cell Starvation: Transcriptome and Proteome Analysis

iTRAQ-based identification of differentially expressed proteins related to growth in the swimming crab, Portunus trituberculatus

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