Multi-omics integration to elucidate the antibacterial mechanism of Streptomyces sp. strain PBSH9.

Exposure to hypoxia disrupts energy metabolism and induces inflammation. However, the pathways and mechanisms underlying energy metabolism disorders caused by hypoxic conditions remain unclear. In the present study, a hypoxic animal model was created and transcriptomic and non-targeted metabolomics techniques were applied to further investigate the pathways and mechanisms of hypoxia exposure that disrupt energy metabolism. Transcriptome results showed that 3,007 genes were significantly differentially expressed under hypoxic exposure, and Gene Ontology annotation analysis and Kyoto Encyclopaedia of Genes and Genomes (KEGG) enrichment analysis showed that the differentially expressed genes (DEGs) were mainly involved in energy metabolism and were significantly enriched in the tricarboxylic acid (TCA) cycle and oxidative phosphorylation (OXPHOS) pathway. The DEGs IDH3A, SUCLA2, and MDH2 in the TCA cycle and the DEGs NDUFA3, NDUFS7, UQCRC1, CYC1 and UQCRFS1 in the OXPHOS pathway were validated using mRNA and protein expression, and the results showed downregulation. The results of non-targeted metabolomics showed that 365 significant differential metabolites were identified under plateau hypoxia stress. KEGG enrichment analysis showed that the differential metabolites were mainly enriched in metabolic processes, such as energy, nucleotide and amino acid metabolism. Hypoxia exposure disrupted the TCA cycle and reduced the synthesis of amino acids and nucleotides by decreasing the concentration of cis-aconitate, α-ketoglutarate, NADH, NADPH and that of most amino acids, purines, and pyrimidines. Bioinformatics analysis was used to identify inflammatory genes related to hypoxia exposure and some of them were selected for verification. It was shown that the mRNA and protein expression levels of IL1B, IL12B, S100A8 and S100A9 in kidney tissues were upregulated under hypoxic exposure. The results suggest that hypoxia exposure inhibits the TCA cycle and the OXPHOS signalling pathway by inhibiting IDH3A, SUCLA2, MDH2, NDUFFA3, NDUFS7, UQCRC1, CYC1 and UQCRFS1, thereby suppressing energy metabolism, inducing amino acid and nucleotide deficiency and promoting inflammation, ultimately leading to kidney damage.

Skeletal muscle fiber types can contribute in part to affecting pork quality parameters. Biceps femoris (Bf) (fast muscle or white muscle) and Soleus (Sol) (slow muscle or red muscle) are two typical skeletal muscles characterized by obvious muscle fiber type differences in pigs. However, the critical proteins and potential regulatory mechanisms regulating porcine skeletal muscle fibers have yet to be clearly defined. In this study, the isobaric Tag for Relative and Absolute Quantification (iTRAQ)-based proteome was used to identify the key proteins affecting the skeletal muscle fiber types with Bf and Sol, by integrating the previous transcriptome data, while function enrichment analysis and a protein–protein interaction (PPI) network were utilized to explore the potential regulatory mechanisms of skeletal muscle fibers. A total of 126 differentially abundant proteins (DAPs) between the Bf and Sol were identified, and 12 genes were found to be overlapping between differentially expressed genes (DEGs) and DAPs, which are the critical proteins regulating the formation of skeletal muscle fibers. Functional enrichment and PPI analysis showed that the DAPs were mainly involved in the skeletal-muscle-associated structural proteins, mitochondria and energy metabolism, tricarboxylic acid cycle, fatty acid metabolism, and kinase activity, suggesting that PPI networks including DAPs are the main regulatory network affecting muscle fiber formation. Overall, these data provide valuable information for understanding the molecular mechanism underlying the formation and conversion of muscle fiber types, and provide potential markers for the evaluation of meat quality.

The development of flower and pollen is a complex biological process that involves multiple metabolic pathways in plants. In revealing novel insights into flower and pollen development underlying male sterility (MS), we conducted an integrated profiling of gene and protein activities in developing buds in cytoplasmic male sterile (CMS) mutants of mustard (Brassica juncea). Using RNA-Seq and label-free quantitative proteomics, 11,832 transcripts and 1780 protein species were identified with significant differential abundance between the male sterile line 09-05A and its maintainer line 09-05B at the tetrad stage and bi-nucleate stage of B. juncea. A large number of differentially expressed genes (DEGs) and differentially abundant proteins (DAPs) involved in carbohydrate and energy metabolism, including starch and sucrose metabolism, tricarboxylic acid (TCA) cycle, glycolysis, and oxidoreductase activity pathways, were significantly downregulated in 09-05A buds. The low expression of these DEGs or functional loss of DAPs, which can lead to an insufficient supply of critical substrates and ATP, could be associated with flower development, pollen development, and changes in fertility in B. juncea. Therefore, this study provided transcriptomic and proteomic information of pollen abortion for B. juncea and a basis for further research on the molecular regulatory mechanism of MS in plants.

The two-line hybrid rice is an important factor of a global crop, but its fertility transition mechanism is unclear. Here, a comparative proteomics and transcriptomics analysis was completed on the two-line hybrid rice line Wuxiang S (WXS) to explore its molecular mechanism and protein regulation during fertility transition. A total of 340 differentially abundant proteins (DAPs) were identified using iTRAQ between the pollen mother cell formation stage (P2) and the meiosis stage (P3). There were 3541 and 4247 differentially expressed genes (DEGs) in P2 and P3 between WXS (Sterile, S)-WXS(S) and WXS (Fertile, F)-WXS(F), respectively, of which 92 and 71 DEGs had corresponding DAPs. Among the DAPs and DEGs, 65 (SP2 vs. FP2) and 55 (SP3 vs. FP3) corresponding DEGs and DAPs (cor-DEGs-DAPs) showed the same expression trend, indicating the cor-DEGs-DAPs genes might play vital roles in WXS fertility transition. Further analysis indicated that cor-DEGs-DAPs proteins were related to energy metabolism-related proteins in anther development and were accompanied by the activation of the stress response pathway and modifications to the cell wall, which ultimately affected the fertility transition of the PTGMS rice line WXS.

The objective of this study was to investigate the key genes and pathways associated with thyroid carcinoma. Based on the microarray data of GSE27155, we identified the differentially expressed genes (DEGs) between four types of thyroid carcinoma samples (papillary carcinoma (PTC), oncocytic carcinoma (OTC), follicular carcinoma (FTC) and anaplastic carcinoma (ATC)) and normal controls. With the obtained DEGs, we performed gene functional interaction (FI) network analysis. Then we conducted Venn diagram analysis to identify the intersection and specific DEGs of the four types of thyroid carcinomas. The intersections DEGs were performed by functional enrichment and transcription factor (TF) prediction analyses. These specific DEGs were performed by pathway enrichment analysis. There were respectively 323, 318, 118 and 1005 DEGs identified in PTC, OTC, FTC and ATC. Twelve sub-network modules were extracted based on gene FI network analysis and eight thyroid carcinoma-associated DEGs were involved in the network, such as TIMP1. Based on the Venn diagram analysis, 27 common DEGs were identified, such as HMGB3 which was regulated by TF of NKX3-1. There were 149 PTC-specific DEGs (like CLDN1), 160 OTC-specific DEGs, 94 FTC-specific DEGs (like PPARG), and 789 ATC-specific DEGs (like CDK1). They were enriched in some pathways, such as Cell cycle, Citrate cycle, and Oxidative phosphorylation. TIMP1, HMGB3, CLDN1, CDK1 and PPARG as well as pathways of Cell cycle, Citrate cycle, and Oxidative phosphorylation may play important roles in the progression of thyroid carcinoma.

BackgroundAnanas comosus var. bracteatus has high ornamental value due to its chimeric leaves. However, the chimeric trait is very unstable in red pineapple plants, and transcriptional variation between the two types of cells (white/green cells) and the molecular mechanism responsible for their albino phenotype remain poorly understood.MethodsComparative transcriptomic and proteomic analyses of the white parts (Whs) and green parts (Grs) of chimeric leaves were performed.ResultsIn total, 1,685 differentially expressed genes (DEGs) (712 upregulated and 973 downregulated) and 1,813 differentially abundant proteins (DAPs) (1,018 with low abundance and 795 with high abundance) were identified. Based on Gene Ontology (Go) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses, the DEGs were mostly involved in carbon fixation in photosynthetic organisms, porphyrin and chlorophyll metabolism and oxidative phosphorylation, while proteomic analysis revealed that DAPs were mostly related to ribosomes, photosynthesis, photosynthesis antennas, and porphyrin and chlorophyll metabolism. Combined analysis showed increased mRNA levels but low abundance of nine proteins level in Whs /Grs related to photosynthetic pigment and photosynthesis. Transcriptional changes, posttranscriptional regulation and translational alterations of key enzymes involved in chlorophyll biosynthesis and photosynthesis may play important roles in the albino parts of chimeric leaves.

BackgroundMaize is one of the most important food crops worldwide. Roots play important role in maize productivity through water and nutrient uptake from the soil. Improving maize root traits for efficient water uptake will help to optimize irrigation and contribute to sustainable maize production. Therefore, we investigated the protein profiles of maize cv. Anyu308 root system divided into Upper root zone (UR), Middle root (MR), and Lower root (LR), by label free quantitative shotgun proteomic approach (LFQ). The aim of our study was to identify proteins and mechanisms associated with enhanced water uptake in different maize root zones under automatic irrigation system.ResultsAt field capacity, MR had the highest water uptake than the UR and LR. We identified a total of 489 differentially abundant proteins (DAPs) by pairwise comparison of MR, LR, and UR. Cluster analysis of DAPs revealed MR and UR had similar protein abundance patterns different from LR. More proteins were differentially abundant in MR/UR compared to LR/MR and LR/UR. Comparisons of protein profiles indicate that the DAPs in MR increased in abundance, compared to UR and LR which had more downregulated DAPs. The abundance patterns, functional category, and pathway enrichment analyses highlight chromatin structure and dynamics, ribosomal structures, polysaccharide metabolism, energy metabolism and transport, induction of water channels, inorganic ion transport, intracellular trafficking, and vesicular transport, and posttranslational modification as primary biological processes related to enhanced root water uptake in maize. Specifically, the abundance of histones, ribosomal proteins, and aquaporins, including mitochondrion electron transport proteins and the TCA cycle, underpinned MR’s enhanced water uptake. Furthermore, proteins involved in folding and vascular transport supported the radial transport of solute across cell membranes in UR and MR. Parallel reaction monitoring analysis was used to confirmed profile of the DAPs obtained by LFQ-based proteomics.ConclusionThe list of differentially abundant proteins identified in MR are interesting candidates for further elucidation of their role in enhanced water uptake in maize root. Overall, the current results provided an insight into the mechanisms of maize root water uptake.

Acinetobacter baumannii has been identified as a critical pathogen, and new antibiotics are urgently needed. Volatile oils, which function as natural antibacterial agents, may provide an effective means of inhibiting A. baumannii. However, the antibacterial activity and mechanism of the volatile oil derived from the dried bark of Cinnamomum cassia (CBV), as well as its additive effect when combined with imipenem (IPM) against A. baumannii, remain unclear. CBV was extracted using the hydrodistillation method and characterized by gas chromatography-mass spectrometry (GC-MS) analysis. The minimum inhibitory concentrations (MICs) of CBV and IPM against A. baumannii were determined using the microdilution method. A checkerboard assay was performed to evaluate the additive effect of CBV (concentration range: 0-1 μL/mL) and IPM (concentration range: 0-256 μg/mL) against A. baumannii, with the fractional inhibitory concentration index (FICI) calculated. A time-kill curve analysis was performed to assess the additive effect of CBV (0.125 μL/mL) and IPM (4 μg/mL) against A. baumannii. Antibiofilm activity was evaluated using a crystal violet staining assay. Cell membrane integrity was assessed using SYTO 9/PI staining based on fluorescence color. Intracellular protein levels were quantified using a BCA kit according to the manufacturer's instructions. Scanning electron microscopy (SEM) was used to observe morphological changes in A. baumannii. Additionally, the antibacterial mechanism was elucidated through a combination of transcriptomic and proteomic analyses. An additive effect (FICI = 0.53) was observed when CBV and IPM were combined against A. baumannii, reducing the MIC of IPM from 256 μg/mL to 4 μg/mL. CBV and IPM inhibited biofilm formation, damaged the cell membrane, and induced intracellular protein leakage in A. baumannii. Compared to CBV or IPM alone, the combination group (at the dosage showing an additive effect) caused significantly greater damage to the cell membrane of A. baumannii. CBV and IPM also induced significant changes at both the transcriptomic and proteomic levels in A. baumannii. Functional analysis revealed that the differentially expressed genes (DEGs) and proteins (DEPs) were involved in multiple pathways. Both CBV and IPM contributed to the observed antibacterial activity. CBV primarily influenced the ribosome pathway, while IPM mainly influenced oxidative phosphorylation. In the combination treatment, the simultaneous targeting of the ribosome and oxidative phosphorylation pathways was identified as the key antibacterial mechanism. This study demonstrated that the combination of CBV and IPM exhibits promising antimicrobial activity against A. baumannii, suggesting that CBV could serve as a potential natural candidate for the development of novel antibiotic agents. While the current findings establish a mechanistic foundation for CBV's antimicrobial effects, further research is necessary to facilitate its clinical translation. Specifically, formulation optimization studies are necessary to enhance the therapeutic viability of the CBV/IPM combination, and comprehensive in vivo investigations are crucial to validate the antibacterial efficacy and safety profile of CBV/IPM prior to clinical application.

Salt stress is one of the most serious abiotic factors that inhibit plant growth. Dunaliella salina has been recognized as a model organism for stress response research due to its high capacity to tolerate extreme salt stress. A proteomic approach based on isobaric tags for relative and absolute quantitation (iTRAQ) was used to analyze the proteome of D. salina during early response to salt stress and identify the differentially abundant proteins (DAPs). A total of 141 DAPs were identified in salt-treated samples, including 75 upregulated and 66 downregulated DAPs after 3 and 24 h of salt stress. DAPs were annotated and classified into gene ontology functional groups. The Kyoto Encyclopedia of Genes and Genomes pathway analysis linked DAPs to tricarboxylic acid cycle, photosynthesis and oxidative phosphorylation. Using search tool for the retrieval of interacting genes (STRING) software, regulatory protein–protein interaction (PPI) networks of the DAPs containing 33 and 52 nodes were built at each time point, which showed that photosynthesis and ATP synthesis were crucial for the modulation of early salinity-responsive pathways. The corresponding transcript levels of five DAPs were quantified by quantitative real-time polymerase chain reaction (qRT-PCR). These results presented an overview of the systematic molecular response to salt stress. This study revealed a complex regulatory mechanism of early salt tolerance in D. salina and potentially contributes to developing strategies to improve stress resilience.

Maize (Zea mays L.) is one of the major staple crops providing human food, animal feed, and raw material support for biofuel production. For its growth and development, maize requires essential macronutrients. In particular, nitrogen (N) plays an important role in determining the final yield and quality of a maize crop. However, the excessive application of N fertilizer is causing serious pollution of land area and water bodies. Therefore, cultivating high-yield and low-N-tolerant maize varieties is crucial for minimizing the nitrate pollution of land and water bodies. Here, based on the analysis of the maize leaf transcriptome and proteome at the grain filling stage, we identified 3957 differentially expressed genes (DEGs) and 329 differentially abundant proteins (DAPs) from the two maize hybrids contrasting in N stress tolerance (low-N-tolerant XY335 and low-N-sensitive HN138) and screened four sets of low-N-responsive genes and proteins through Venn diagram analysis. We identified 761 DEGs (253 up- and 508 down-regulated) specific to XY335, whereas 259 DEGs (198 up- and 61 down-regulated) were specific to HN138, and 59 DEGs (41 up- and 18 down-regulated) were shared between the two cultivars under low-N-stress conditions. Meanwhile, among the low-N-responsive DAPs, thirty were unique to XY335, thirty were specific to HN138, and three DAPs were shared between the two cultivars under low-N treatment. Key among those genes/proteins were leucine-rich repeat protein, DEAD-box ATP-dependent RNA helicase family proteins, copper transport protein, and photosynthesis-related proteins. These genes/proteins were involved in the MAPK signaling pathway, regulating membrane lipid peroxidation, and photosynthesis. Our results may suggest that XY335 better tolerates low-N stress than HN138, possibly through robust low-N-stress sensing and signaling, amplified protein phosphorylation and stress response, and increased photosynthesis efficiency, as well as the down-regulation of 'lavish' or redundant proteins to minimize N demand. Additionally, we screened glutathione transferase 42 (ZmGST42) and performed physiological and biochemical characterizations of the wild-type (B73) and gst42 mutant at the seedling stage. Resultantly, the wild-type exhibited stronger tolerance to low N than the mutant line. Our findings provide a better understanding of the molecular mechanisms underlying low-N tolerance during the maize grain filling stage and reveal key candidate genes for low-N-tolerance breeding in maize.

Soil cadmium pollution has increasingly become a serious problem for crop production, which drastically attenuates plant growth and food safety. Although N6-methyladenosine (m6A) methylation is crucial for plant response to various stresses, the regulatory mechanism underlying m6A modification during cadmium (Cd) stress remains unclear. This study investigated the physiological responses, transcriptome-wide m6A methylome, and proteome changes in tomato roots exposed to 50 μmol · L−1 CdCl2. Excess Cd restricted plant growth, altered the antioxidant system and disrupted mineral nutrient absorption. We identified a negative correlation between m6A levels and gene transcription for 150 out of 198 differentially expressed genes (DEGs) that were hypomethylated but mRNA up-regulated. Cd stress also enhanced translational efficiency, particularly for differentially abundant proteins (DAPs). Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that differentially m6A modified genes (DMGs), DEGs, and DAPs were commonly enriched in phenylpropanoid biosynthesis, glutathione metabolism, and ABC transporters, reflecting cell wall barriers, chelation, and transport of Cd, respectively. Finally, we confirmed the Cd-transport activity of eight putative metal transporters identified in DMGs, DEGs, or DAPs by yeast complementaion experiments, and pharmacologically investigated the effect of m6A modification on their expression. Treatment with the m6A methylation inhibitor 3-deazaneplanocin A (3-DA) reduced SlIRT1/2 expression and increased SlNRAMP3/SlZIP4 expression, while the m6A demethylase inhibitor meclofenamic acid (MA) treatment decreased SlNRAMP3 expression but elevated SlIRT2 expression under Cd stress. Our findings provide novel insights into the interplay between m6A modification, transcription, and translation under Cd stress and the associated plant stress response.

Subcutaneous fat and intramuscular fat (IMF) deposition are closely related to meat production and pork quality. Dingyuan pig, as a native pig breed in China, low selection leads to obvious genetic and phenotypic differences in the population. Individuals with extreme fat content in the population are ideal models for studying the mechanism of fat deposition. In this study, we used RNA-Seq and tandem mass tags-based (TMT) proteomics to analyze the key pathways and genes that specifically regulate subcutaneous fat and IMF deposition in Dingyuan pigs. We identified 191 differentially expressed genes (DEGs) and 61 differentially abundant proteins (DAPs) in the high and low back fat thickness (HBF, LBF) groups, 85 DEGs and 12 DAPs were obtained in the high and low intramuscular fat (HIMF, LIMF) groups. The functional analysis showed that the DEGs and DAPs in the backfat groups were mainly involved in carbohydrates, amino acids, and fatty acids metabolism, whereas the IMF groups were involved in the insulin pathway, longevity, and some disease-related pathways. We found 40 candidate genes that might tissue-specifically lipids deposition for subcutaneous and intramuscular fat. Our research provides theoretical reference materials for the improvement of fat deposition traits of local pig breeds in my country.

Cytoplasmic male sterility (CMS) where no functional pollen is produced has important roles in wheat breeding. The anther is a unique organ for male gametogenesis and its abnormal development can cause male sterility. However, the mechanisms and regulatory networks related to plant male sterility are poorly understood. In this study, we conducted comparative analyses using isobaric tags for relative and absolute quantification (iTRAQ) of the pollen proteins in a CMS line and its wheat maintainer. Differentially abundant proteins (DAPs) were analyzed based on Gene Ontology classifications, metabolic pathways and transcriptional regulation networks using Blast2GO. We identified 5570 proteins based on 23,277 peptides, which matched with 73,688 spectra, including proteins in key pathways such as glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase and 6-phosphofructokinase 1 in the glycolysis pathway, isocitrate dehydrogenase and citrate synthase in the tricarboxylic acid cycle and nicotinamide adenine dinucleotide (NADH)-dehydrogenase and adenosine-triphosphate (ATP) synthases in the oxidative phosphorylation pathway. These proteins may comprise a network that regulates male sterility in wheat. Quantitative real time polymerase chain reaction (qRT-PCR) analysis, ATP assays and total sugar assays validated the iTRAQ results. These DAPs could be associated with abnormal pollen grain formation and male sterility. Our findings provide insights into the molecular mechanism related to male sterility in wheat.

The freezing tolerance of roots is crucial for winter turnip rape (Brassica rapa L.) survival in the winter in Northwest China. Cold acclimation (CA) can alleviate the root damage caused by freezing stress. To acknowledge the molecular mechanisms of freezing tolerance in winter turnip rape, two Brassica rapa genotypes, freezing stressed after the induction of cold acclimation, were used to compare the proteomic profiles of roots by isobaric tags for relative and absolute quantification (iTRAQ). Under freezing stress (−4 °C) for 8 h, 139 and 96 differentially abundant proteins (DAPs) were identified in the roots of “Longyou7” (freezing-tolerant) and “Tianyou4” (freezing-sensitive), respectively. Among these DAPs, 91 and 48 proteins were up- and down-accumulated in “Longyou7”, respectively, and 46 and 50 proteins were up- and down-accumulated in “Tianyou4”, respectively. Under freezing stress, 174 DAPs of two varieties were identified, including 9 proteins related to ribosome, 19 DAPs related to the biosynthesis of secondary metabolites (e.g., phenylpropanoid and the lignin pathway), and 22 down-accumulated DAPs enriched in oxidative phosphorylation, the pentose phosphate pathway, fructose and mannose metabolism, alpha-linolenic acid metabolism, carbon fixation in photosynthetic organisms, ascorbate and aldarate metabolism. The expressional pattern of the genes encoding the 15 significant DAPs were consistent with the iTRAQ data. This work indicates that protein biosynthesis, lignin synthesis, the reduction of energy consumption and a higher linolenic acid content contribute to the freezing tolerance of winter turnip rape. Functional analyses of these DAPs would be helpful in dissecting the molecular mechanisms of the stress responses in B. rapa.

BackgroundDrought stress restricts the growth, distribution and productivity of alfalfa (Medicago sativa L.). In order to study the response differences of alfalfa cultivars to drought stress, we previously carried out physiological and molecular comparative analysis on two alfalfa varieties with contrasting drought resistance (relatively drought-tolerant Longdong and drought-sensitive Algonquin). However, the differences in proteomic factors of the two varieties in response to drought stress still need to be further studied. Therefore, TMT-based quantitative proteomic analysis was performed using leaf tissues of the two alfalfa cultivars to identify and uncover differentially abundant proteins (DAPs).ResultsIn total, 677 DAPs were identified in Algonquin and 277 in Longdong under drought stress. Subsequently, we conducted various bioinformatics analysis on these DAPs, including subcellular location, functional classification and biological pathway enrichment. The first two main COG functional categories of DAPs in both alfalfa varieties after drought stress were ‘Translation, ribosomal structure and biogenesis’ and ‘Posttranslational modification, protein turnover, chaperones’. According to KEGG database, the DAPs of the two alfalfa cultivars after drought treatment were differentially enriched in different biological pathways. The DAPs from Algonquin were enriched in ‘photosynthesis’ and ‘ribosome’. The pathways of ‘linoleic acid metabolism’, ‘protein processing in endoplasmic reticulum’ and ‘RNA transport’ in Longdong were significantly enriched. Finally, we found significant differences in DAP enrichment and expression patterns between Longdong and Algonquin in glycolysis/glycogenesis, TCA cycle, photosynthesis, protein biosynthesis, flavonoid and isoflavonoid biosynthesis, and plant-pathogen interaction pathway after drought treatment.ConclusionsThe differences of DAPs involved in various metabolic pathways may explain the differences in the resistance of the two varieties to drought stress. These DAPs can be used as candidate proteins for molecular breeding of alfalfa to cultivate new germplasm with more drought tolerance to adapt to unfavorable environments.