Transport-related Proteins Research Articles

The SNARE protein LbSYP61 participates in salt secretion in Limonium bicolor

Soil salinization is a global problem that inhibits plant growth and limits crop production. Recretohalophyte plants secrete excess salt via salt glands, thus avoiding excessive accumulation of salt within their cells and protecting themselves from salt-associated damage. However, the molecular mechanisms underlying salt secretion via salt glands are unclear. Here, we used brefeldin A (BFA), a specific inhibitor of Golgi-mediated cell secretion, to test whether vesicle transport participates in salt secretion in the recretohalophyte Limonium bicolor using leaf disc and non-invasive micrometric assays. Indeed, salt secretion via salt glands was significantly inhibited, the Golgi apparatus in the salt gland was damaged, and acidic phosphatase activity was significantly inhibited following a 200 μg/ml BFA treatment. To further examine the role of vesicle transport in salt secretion, we used virus-induced gene silencing (VIGS) to downregulate the gene encoding a trans Golgi-localized soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein, LbSYP61. After silencing LbSYP61, the ability of L. bicolor to secrete salt via salt glands was also significantly reduced. These results provide direct evidence that vesicular transport participates in salt secretion via salt glands and the vesicular transport-related protein LbSYP61 plays an important role in this process.

Analyzing the grape leaf proteome and photosynthetic process provides insights into the injury mechanisms of ozone stress

Ozone can cause damage to plant tissues. However, little information is available about this global and systematic damage mechanism of grape leaf photosynthetic apparatus. In this study, tandem mass tags (TMT) analysis and physiological responses indicated severe damage to the chloroplasts, with differentially expressed proteins mainly distributed in the chloroplasts. Under ozone stress, the expression of the PSI and PSII structural proteins PsbR, PsaK, PsaA and PSII reactive center protein D1 levels were down-regulated, and the inhibition of D1 protein turnover further aggravated the photodamage of photosystem II (PSII), resulting in a significant decrease in the actual photochemical efficiency of PSII (Y(II)) and actual photochemical efficiency of PSI (Y(I)). The xanthophyll cycle-related protein VDE and ZEP and the NDH subunit of the cyclic electron transport-related protein PGR5-like protein 1A and subcomplex B1 protein were upregulated in response to ozone stress, which was accompanied by a high level of ETRI-ETRII ratio. However, these defense mechanisms are not strong enough. Several dysfunctional antioxidant-related proteins have shown limited ability to scavenge reactive oxygen species, leading to reactive oxygen species burst and eventual photodamage. The system analysis provides new insights into the damage and molecular defense mechanisms of photosynthetic apparatus by ozone.

The Effect of Silibinin on Protein Expression Profile in White Adipose Tissue of Obese Mice.

ObjectiveTo investigate the effect of silibinin on the protein expression profile of white adipose tissue (WAT) in obese mice by using Tandem Mass Tag (TMT) and liquid chromatography-tandem mass spectrometry (LC-MS/MS).MethodsAccording to experimental requirements, 36 C57BL/6JC mice were randomly divided into normal diet group (WC group), high fat diet group (WF group), and high fat diet + silibinin group (WS group). WS group was intragastrically administered with 54 mg/kg body weight of silibinin, and the WC group and the WF group were intragastrically administered with equal volume of normal saline. Serum samples were collected to detect fasting blood glucose and blood lipids. IPGTT was used to measure the blood glucose value at each time point and calculate the area under the glucose curve. TMT combined with LC-MS/MS were used to study the expression of WAT, and its cellular processes, biological processes, corresponding molecular functions, and related network molecular mechanisms were analyzed by bioinformatics. Finally, RT-PCR and LC-MS/MS were used to detect the mRNA and protein expressions of FABP5, Plin4, GPD1, and AGPAT2, respectively.ResultsAlthough silibinin did not reduce the mice's weight, it did improve glucose metabolism. In addition, silibinin decreased the concentration of TC, TG, and LDL-C and increased the concentration of HDL-C in the serum of mice. In the WF/WS group, 182 differentially expressed proteins were up-regulated and 159 were down-regulated. While in the WS/WF group, 362 differentially expressed proteins were up-regulated and 176 were down-regulated. Further analysis found that these differential proteins are mainly distributed in the peroxisome proliferation-activated receptor (PPAR), lipolysis of fat cells, metabolism of glycerides, oxidative phosphorylation, and other signaling pathways, and participate in cell processes and lipid metabolism through catalysis and integration functions. Specifically, silibinin reduced the expression of several key factors such as FABP5, Plin4, GPD1, and AGPTA2.ConclusionHigh fat diet (HFD) can increase the expression of lipid synthesis and transport-related proteins and reduce mitochondrial related proteins, thereby increasing lipid synthesis, reducing energy consumption, and improving lipid metabolism in vivo. Silibinin can reduce lipid synthesis, increase energy consumption, and improve lipid metabolism in mice in vivo.

Substrate specificity of plastid phosphate transporters in a non-photosynthetic diatom and its implication in evolution of red alga-derived complex plastids

The triose phosphate transporter (TPT) is one of the prerequisites to exchange metabolites between the cytosol and plastids. In this study, we demonstrated that the four plastid TPT homologues in the non-photosynthetic diatom Nitzschia sp. NIES-3581 were highly likely integrated into plastid envelope membranes similar to counterparts in the model photosynthetic diatom Phaeodactylum tricornutum, in terms of target membranes and C-terminal orientations. Three of the four Nitzschia TPT homologues are capable of transporting various metabolites into proteo-liposomes including triose phosphates (TPs) and phosphoenolpyruvate (PEP), the transport substrates sufficient to support the metabolic pathways retained in the non-photosynthetic diatom plastid. Phylogenetic analysis of TPTs and closely related transporter proteins indicated that diatoms and other algae with red alga-derived complex plastids possess only TPT homologues but lack homologues of the glucose 6-phosphate transporter (GPT), xylulose 5-phosphate transporter (XPT), and phosphoenolpyruvate transporter (PPT). Comparative sequence analysis suggests that many TPT homologues of red alga-derived complex plastids potentially have the ability to transport mainly TPs and PEP. TPTs transporting both TPs and PEP highly likely mediate a metabolic crosstalk between a red alga-derived complex plastid and the cytosol in photosynthetic and non-photosynthetic species, which explains the lack of PPTs in all the lineages with red alga-derived complex plastids. The PEP-transporting TPTs might have emerged in an early phase of endosymbiosis between a red alga and a eukaryote host, given the broad distribution of that type of transporters in all branches of red alga-derived complex plastid-bearing lineages, and have probably played a key role in the establishment and retention of a controllable, intracellular metabolic connection in those organisms.

Characterization and immunostimulatory activity of extracellular vesicles from Filifactor alocis.

Filifactor alocis, a gram-positive, obligate anaerobic rod, is an emerging periodontal pathogen that is frequently isolated from patients with periodontitis, peri-implantitis, and apical periodontitis. Recent studies have shown that extracellular vesicles (EVs) from gram-negative periodontal pathogens, so-called outer membrane vesicles (OMVs), harbor various effector molecules responsible for inducing host inflammatory responses. However, there are no reports of EVs from F. alocis. In this study, we purified and characterized the protein profiles of EVs from F. alocis and investigated their immunostimulatory activity on human monocytic THP-1 and human oral keratinocyte HOK-16B cell lines. Highly pure EVs were obtained from F. alocis using density gradient ultracentrifugation. Nanoparticle tracking analysis and transmission electron microscopy showed that F. alocis EVs were between 50 and 270nm in diameter. Proteome analysis identified 28 proteins, including lipoproteins, autolysins, F. alocis complement inhibitor (FACIN), transporter-related proteins, metabolism-related proteins, and ribosomal proteins. Human cytokine array analysis showed that F. alocis EVs remarkably induced the expression of CCL1, CCL2, MIP-1, CCL5, CXCL1, CXCL10, ICAM-1, IL-1β, IL-1ra, IL-6, IL-8, MIF, SerpinE, and TNF-α in THP-1 cells and CXCL1, G-CSF, GM-CSF, IL-6, and IL-8 in HOK-16B cells. The immunostimulatory activity of F. alocis EVs was similar to that of the whole bacterial cells. Our findings provide new insight into the role of EVs from gram-positive oral bacteria in periodontal diseases.

SANS (USH1G) Molecularly Links the Human Usher Syndrome Protein Network to the Intraflagellar Transport Module by Direct Binding to IFT-B Proteins.

The human Usher syndrome (USH) is a retinal ciliopathy, characterized by profound congenital deafness, variable vestibular dysfunction and pre-pubertal onset of retinitis pigmentosa. In the effected sensory cells, USH protein networks are assumed to function in ciliary transport processes. The USH1G protein SANS is a scaffold of the ciliary/periciliary USH protein network of photoreceptor cells. Moreover, SANS is associated with microtubules, the transport routes for protein delivery toward the cilium. To enlighten the role of SANS in ciliary transport processes, we aimed to identify transport related proteins associated with SANS. The intraflagellar transport (IFT) system is a conserved mechanism for bi-directional transport toward and through primary cilia. Thus, we tested the direct binding of SANS to IFT molecules, namely IFT20, IFT57, and IFT74 in 1:1 yeast-two-hybrid assay. The identified SANS-IFT interactions were validated in vitro via independent complementary interaction assays and in cells by applying membrane targeting assays. Quantitative immunofluorescence microscopy revealed the co-localization of SANS with IFT20, IFT52, and IFT57 particularly at ciliary base of wild type mouse photoreceptor cells. Analysis of photoreceptor cells of SANS knock out mice revealed the decrease of IFTs in the ciliary compartment indicating a role of SANS in the proper positioning of IFT-B molecules in primary cilia. Our study demonstrated direct binding of IFT complex B proteins IFT52 and IFT57 to the N-terminal ankyrin repeats and the central domain of SANS. Our data also indicate that pathologic mutations in the N-terminus of SANS lead to the loos of SANS binding to IFT-B molecules. Our findings provide direct evidence for a molecular link between the ciliary USH protein network and the IFT transport module in primary cilia.

Developmental changes in transporter and receptor protein expression levels at the rat blood-brain barrier based on quantitative targeted absolute proteomics

The blood-brain barrier (BBB) transport systems regulate the supply of nutrients, amino acids, vitamins, and hormones to the developing brain, as well as blocking the entry of xenobiotics and drugs. The purpose of this study was to clarify the developmental changes in the absolute protein expression levels of BBB transport-related proteins in developing rat brain capillaries, using quantitative targeted absolute proteomics (QTAP). The changing patterns of ATP-binding cassette (ABC) and solute carrier (SLC) transporters, receptors, and tight junction/adherence junction-related proteins were classified into 4 types: uphill (continuously increasing expression from postnatal day (P) 1 to P56), bell-shape/inverted bell-shape (increased/decreased expression from P1 to P14 followed by decreased/increased expression from P21 to P56), downhill (continuously decreasing expression from P1 to P56), and constant (no significant difference from P1 to P56). Proteins showing uphill-type expression included P-glycoprotein/Mdr1a/Abcb1, Mrp4/Abcc4, Bcrp/Abcg2, Glut1/Slc2a1, Oatp1c1/Slco1c1, FcRn, 4F2hc/Slc3a2, claudin-5, caveolin-1, Cd29/integrin β1. Those showing bell-shape/inverted bell-shape expression included Mct1/Slc16a1, Oat3/Slc22a8, Tfr1, Lrp1, and CD147. On the other hand, Cat1/Slc7a1 and Cd54/Icam-1 showed downhill expression, and Insr showed constant expression. These results suggest that the protein expression levels of transporters and receptors at the BBB change in various ways to meet the changing requirements of the developing brain.

Enhancing the retrograde axonal transport by curcumin promotes autophagic flux in N2a/APP695swe cells.

The accumulation of autophagosomes and dysfunction at the axonal terminal of neurons play crucial roles in the genesis and development of Alzheimer’s disease (AD). Abnormalities in neuron axonal transport-related proteins prevent autophagosome maturation in AD. Curcumin, a polyphenol plant compound, has been shown to exert neuroprotective effects by increasing autophagy in AD, but the underlying mechanism of its effect on autophagy axon transport remains elusive. This study investigated the effects of curcumin on autophagosome formation and axonal transport in N2a/APP695swe cells (AD cell model) as well as the mechanism underlying those effects. Curcumin treatment significantly increased the expression of Beclin1, Atg5, and Atg16L1, induced the formation of autophagosomes, and promoted autophagosome–lysosome fusion in N2a/APP695swe cells. At the same time, curcumin promoted the expression of dynein, dynactin, and BICD2 as well as their binding to form the retrograde axonal transport molecular motor complex. Moreover, curcumin also increased the expression of the scaffolding proteins Rab7- interacting lysosomal protein (RILP) and huntingtin in N2a/APP695swe cells. Taken together, our findings indicate that curcumin increases autophagic flux by promoting interactions among autophagic axonal transport-related proteins and inducing lysosome–autophagosome fusion. This study provides evidence suggesting the potential use of curcumin as a novel treatment for AD.

Chronic cerebral hypoperfusion alters amyloid-β transport related proteins in the cortical blood vessels of Alzheimer’s disease model mouse

Abnormal accumulation of amyloid-β (Aβ) peptide defines progression of Alzheimer’s disease (AD) pathology in brain. Here, we investigated expressive changes of two main Aβ transport receptors low-density lipoprotein receptor related protein-1 (LRP1) and receptor for advanced glycation end products (RAGE) in a novel AD mice (APP23) with chronic cerebral hypoperfusion (CCH) model, moreover, examined a protective effect of a free radical scavenger edaravone (Eda). In contrast to wild type (WT) and APP23 mice, CCH strongly accelerated abnormal Aβ40 depositions and cerebral amyloid angiopathy (CAA) pathology, increased both LRP1 and RAGE expressions in brain parenchyma, while a decrease of LRP1 and an increase of RAGE were observed in vascular endothelial cells at age 12 months (M) of AD mice. Furthermore, CCH strongly increased expressions of two hypoxia-related proteins hypoxia inducible factor-1α (HIF-1α) and heme oxygenase-1 (HO-1), two oxidative-related proteins 4-hydroxy-2-nonenal (4-HNE) and 8-hydroxy-2′-deoxyguanosine (8-OHdG), and decreased both two vital nutrient transporter proteins major facilitator super family domain containing 2a (Mfsd2a) and glucose transporter 1 (Glut1) expressions. Such the above abnormal pathological changes were significantly ameliorated by edaravone treatment. The present study demonstrated that CCH strongly enhanced primary AD pathology causing double imbalances of Aβ efflux and influx transport related proteins in the cortical blood vessels in AD mice, and that such a neuropathologic abnormality was greatly ameliorated by Eda.

A Global Proteomic Change in Petroleum Hydrocarbon-Degrading Pseudomonas aeruginosa in Response to High and Low Concentrations of Petroleum Hydrocarbons.

In this study, iTRAQ analysis and bioinformatics analysis were used to reveal the changes in key proteins induced by different concentrations of petroleum hydrocarbons during the biodegradation of petroleum hydrocarbons in Pseudomonas aeruginosa P6. Sixty-three proteins were identified as differentially expressed proteins, and all of them were strongly associated with the cellular processes related to the biodegradation of petroleum hydrocarbons. The results further showed that among the differentially expressed proteins, 3 chemotaxis-related proteins, 10 terminal oxidation of short-chain alkane-related proteins, and 13 transmembrane transport-related proteins were down regulated, while 1 uptake of petroleum hydrocarbon-related protein, 3 terminal oxidation of long-chain alkane-related proteins, 4 dehydrogenation-related proteins, 12 β-oxidation-related proteins, and 2 metabolisms of acyl-CoA-related proteins were up regulated. These results indicated that during the biodegradation of petroleum hydrocarbons in P. aeruginosa P6, the activity of chemotaxis, the terminal oxidation of short-chain alkanes, and transmembrane transport decreased, while the activity of the uptake of petroleum hydrocarbons, the terminal oxidation of long-chain alkanes, dehydrogenation, β-oxidation, and the metabolism of acyl-CoA increased under the 20,000mg/L petroleum hydrocarbon condition compared with the 500mg/L petroleum hydrocarbon condition. The findings revealed changes in the key proteins and the corresponding cellular process of the biodegradation of petroleum hydrocarbons in P. aeruginosa P6 under high and low concentrations of petroleum hydrocarbons and provided references for future studies.

Evaluation of dynamic developmental processes and the molecular basis of the high body fat percentage of different proglottid types of Moniezia expansa

BackgroundMoniezia expansa (Cyclophyllidea: Anoplocephalidae) is a large species of tapeworm that occurs in sheep and cattle and inhabits the small intestine, causing diarrhea and weight declines, leading to stockbreeding losses. Interestingly, the body fat percentage of M. expansa, which lacks the ability to synthesize fatty acids, is as high as 78% (dry weight) and all of the proglottids of M. expansa exhibit a dynamic developmental process from top to bottom. The aim of this paper is to identify the molecular basis of this high body fat percentage, the dynamic expression of developmental genes and their expression regulation patterns.ResultsFrom 12 different proglottids (four sections: scolex and neck, immature, mature and gravid with three replicates), 13,874 transcripts and 680 differentially expressed genes (DEGs) were obtained. The gene expression patterns of the scolex and neck and immature proglottids were very similar, while those of the mature and gravid proglottids differed greatly. In addition, 13 lipid transport-related proteins were found in the DEGs, and the expression levels showed an increasing trend in the four proglottid types. Furthermore, it was shown that 33 homeobox genes, 9 of which were DEGs, had the highest expression in the scolex and neck section. The functional enrichment results of the DEGs were predominantly indicative of development-related processes, and there were also some signal transduction and metabolism results. The most striking result was the finding of Wnt signaling pathways, which appeared multiple times. Furthermore, the weighted gene co-expression networks were divided into 12 modules, of which the brown module was enriched with many development-related genes.ConclusionsWe hypothesize that M. expansa uses lipid transport-associated proteins to transport lipids from the host gut to obtain energy to facilitate its high fecundity. In addition, homeobox genes and Wnt signaling pathways play a core role in development and regeneration. The results promote research on the cell differentiation involved in the continuous growth and extension of body structures.

Comprehensive Analysis of Soybean Mosaic Virus P3 Protein Interactors and Hypersensitive Response-Like Lesion-Inducing Protein Function

Soybean mosaic virus (SMV) is one of the most prevalent and important pathogens of soybean, which produces 11 proteins, and the third protein, P3, was suggested to be involved in virus movement and replication, as well as host infection. During the virus infection, host proteins are essential in the virus cycle. However, there is no comprehensive report on the network of host proteins that interact with P3. Fifty-one interactors were identified by using the P3 protein as the bait against the SMV SC15 strain-challenged soybean cDNA library. These proteins were classified into five groups, including transport and protein transport-related proteins, defense and disease-related proteins, photosynthesis proteins, cellular metabolic proteins, and unknown proteins. Among these proteins, the protein defined as hypersensitive response-like lesion-inducing (HRLI) appeared multiple times and showed strong affinity with P3, which indicated its important role in SMV infection. Thus, it was chosen for further investigation. Phylogenetic classification showed that paralog proteins GmHRLI-1 and GmHRLI-2 clustered together and shared 90% homologous identity. Bimolecular fluorescence complementation (BiFC) assay was carried out to confirm the interaction, and fluorescence was detected at the cell periplasmic as well as at the nucleus. Subcellular localization showed that GmHRLI was localized to the cell periplasmic, while the co-localization of GmHRLI and P3 signals was also observed in the nucleus, suggesting that GmHRLI could interact with P3 and promoted the translation of P3 to the nucleus. Moreover, the gene expression of GmHRLI was abundant in the roots, leaves, and flowers, and could be induced by SMV infection, suggesting its involvement in SMV infection. Our results together lay the foundation to explore the mechanisms of P3 in the HR process and the HRLI protein function in SMV response.

Midrib Sucrose Accumulation and Sugar Transporter Gene Expression in YCS-Affected Sugarcane Leaves

Sucrose accumulation and decreased photosynthesis are early symptoms of yellow canopy syndrome (YCS) in sugarcane (Saccharum spp.), and precede the visual yellowing of the leaves. To investigate broad-scale gene expression changes during YCS-onset, transcriptome analyses coupled to metabolome analyses were performed. Across leaf tissues, the greatest number of differentially expressed genes related to the chloroplast, and the metabolic processes relating to nitrogen and carbohydrates. Five genes represented 90% of the TPM (Transcripts Per Million) associated with the downregulation of transcription during YCS-onset, which included PSII D1 (PsbA). This differential expression was consistent with a feedback regulatory effect upon photosynthesis. Broad-scale gene expression analyses did not reveal a cause for leaf sugar accumulation during YCS-onset. Interestingly, the midrib showed the greatest accumulation of sugars, followed by symptomatic lamina. To investigate if phloem loading/reloading may be compromised on a gene expression level – to lead to leaf sucrose accumulation - sucrose transport-related proteins of SWEETs, Sucrose Transporters (SUTs), H+-ATPases and H+-pyrophosphatases (H+-PPases) were characterised from a sugarcane transcriptome and expression analysed. Two clusters of Type I H+-PPases, with one upregulated and the other downregulated, were evident. Although less pronounced, a similar pattern of change was observed for the H+-ATPases. The disaccharide transporting SWEETs were downregulated after visual symptoms were present, and a monosaccharide transporting SWEET upregulated preceding, as well as after, symptom development. SUT gene expression was the least responsive to YCS development. The results are consistent with a reduction of photoassimilate movement through the phloem leading to sucrose build-up in the leaf.

Preventive Effects of Fucoidan and Fucoxanthin on Hyperuricemic Rats Induced by Potassium Oxonate.

The purpose of this study was to investigate the preventive effects of fucoidan (Fc) and fucoxanthin (Fx) on hyperuricemic rats. Sprague Dawley (SD) rats were randomly assigned to seven groups: a control group, a hyperuricemia (HUA) group, low- and high-dose Fx groups, a Fc group, a combination Fc and Fx group, and a positive control group. Three weeks after the interventions, each group was given potassium oxonate (PO) and hypoxanthine (HX) to induce HUA in all groups except for the control group, and the rats were then sacrificed. Blood and urine were analyzed for biochemical properties, and differences in urine volume were determined. Livers and kidneys were collected to analyze xanthine oxidase (XO) activity and the expression of uric acid (UA) transporter-related proteins (GLUT9, ABCG2, OAT1, URAT1). The results show that HUA was successfully induced by PO/HX after 4 h of administration. The activity of XO was significantly reduced by a combination of Fc and Fx. In the combination group, both ABCG2 and OAT1 increased significantly, whereas GLUT9 and URAT1 decreased significantly. In summary, the combination of Fc and Fx can inhibit the activity of XO in the liver and regulate the expression of proteins related to UA transporter in the kidney to reduce the UA level in serum.

Copper accumulation, subcellular partitioning and physiological and molecular responses in relation to different copper tolerance in apple rootstocks.

To unravel the physiological and molecular regulation mechanisms underlying the variation in copper (Cu)accumulation, translocation and tolerance among five apple rootstocks, seedlings were exposed to either basal or excess Cu. Excess Cu suppressed plant biomass and root architecture, which was less pronounced in Malus prunifolia Borkh., indicating its relatively higher Cu tolerance. Among the five apple rootstocks, M. prunifolia exhibited the highest Cu concentration and bio-concentration factor in roots but the lowest translocation factor, indicating its greater ability to immobilize Cu and restrict translocation to the aerial parts. Higher Cu concentration in cell wall fraction but lower Cu proportion in membrane-containing and organelle-rich fractions were found in M. prunifolia. Compared with the other four apple rootstocks under excess Cu conditions, M. prunifolia had a lower increment of hydrogen peroxide in roots and leaves and malondialdehyde in roots, but higher concentrations of carbohydrates and enhanced antioxidants. Transcript levels of genes involved in Cu uptake, transport and detoxification revealed species-specific differences that are probably related to alterations in Cu tolerance. M. prunifolia had relatively higher gene transcript levels including copper transporters 2 (COPT2), COPT6 and zinc/iron-regulated transporter-related protein 2 (ZIP2), which probably took part in Cu uptake, and C-type ATP-binding cassette transporter 2 (ABCC2), copper chaperone for Cu/Zn superoxide dismutase (CCS), Cu/Zn superoxide dismutase 1 (CSD1) and metallothionein 2 (MT2) probably implicated in Cu detoxification, and relatively lower mRNA levels of yellow stripe-like transporter 3 (YSL3) and heavy metal ATPase 5 (HMA5) involved in transport of Cu to aerial parts. These results suggest that M. prunifolia is more tolerant to excess Cu than the other four apple rootstocks under the current experimental conditions, which is probably attributed to more Cu retention in roots, subcellular partitioning, well-coordinated antioxidant defense mechanisms and transcriptional expression of genes involved in Cu uptake, translocation and detoxification.

Genome-Wide Association Study Reveals Novel Genomic Regions Associated With High Grain Protein Content in Wheat Lines Derived From Wild Emmer Wheat.

Grain protein content (GPC) and yield are of two important traits in wheat, but their negative correlation has hampered their simultaneous improvement in conventional breeding. Wild emmer wheat (Triticum turgidum ssp. dicoccoides) is an important genetic resource for wheat quality improvement. In this study, we report a genome-wide association study (GWAS) using 13116 DArT-seq markers to characterize GPC in 161 wheat lines derived from wild emmer. Using a general linear model, we identified 141 markers that were significantly associated with GPC, and grouped into 48 QTL regions. Using both general linear model and mixed linear model, we identified four significant markers that were grouped into two novel QTL regions on chromosomes 2BS (QGpc.cd1-2B.1) and 7BL (QGpc.cd1-7B.2). The two QTLs have no negative effects on thousand kernel weight (TKW) and should be useful for simultaneous improvement of GPC and TKW in wheat breeding. Searches of public databases revealed 61 putative candidate/flanking genes related to GPC. The putative proteins of interest were grouped in four main categories: enzymes, kinase proteins, metal transport-related proteins, and disease resistance proteins. The linked markers and associated candidate genes provide essential information for cloning genes related to high GPC and performing marker-assisted breeding in wheat.

NcRNAs associated with drug resistance and the therapy of digestive system neoplasms.

Digestive system cancer remains a common cancerand the main cause of cancer-related death worldwide. Drug resistance is a major challenge in the therapy of digestive system cancer, andrepresents a primary obstaclein the treatment of cancer by restricting the efficiency of both traditional chemotherapy and biological therapies. Existing studiesindicatethat noncoding RNAs play an important role in the evolution and progression of drug resistance in digestive system cancer, mainly by modulating drug transporter-related proteins, DNA damage repair,cell-cycle-related proteins, cell apoptosis-related proteins, drug target-related proteins, and the tumor microenvironment. In this review, we address the potential mechanisms of ncRNAs underlying drug resistance in digestive system tumors and discuss the possible application of ncRNAs against drug resistance in digestive system tumors.

Influence of organic substrates on nutrient accumulation and proteome changes in tomato-roots

The gradually accumulation of nutrients in organic substrates during growing season may have adverse effects on plant development. This study was conducted to determine the mode of nutrient accumulation and alteration at molecular level induced by the excess nutrient stress in tomato. The ICP-MS was used to analyze mineral ions in root zone solution and nutrients uptake by tomato plants; and iTRAQ-based proteomic analysis was conducted to identify proteome changes in tomato roots. Coir and peat-vermiculite were used in substrate treated experiments. Results showed that the content of K+ was higher in coir than in peat-vermiculite whereas it was the opposite for Ca2+. The concentrations of NO3−, SO42- and Mg2+ were significantly higher in peat-vermiculite and coir substrates than in water culture. The peat-vermiculite substrates generally enhanced Ca uptake but reduced P uptake by plants, when compared to both coir and water culture. Compared to water culture, functional annotation analysis of the root proteome revealed that the excess nutrient accumulation induced complex proteomic alterations involved in mineral ion binding and transport. A total of 358 differentially abundant proteins (DAPS) were identified, including 11 mineral ion binding and transport related proteins, such as calmodulin-like protein and nitrate transporter 3.2 under peat-vermiculite and coir cultivations. RT-qPCR was used to validate nine genes encoding DAPS. We believe that these indicators will contribute to a better control of soilless culture systems and a waste reduction in production of tomatoes.

Effect of <italic>hfq2</italic> gene in <italic>Aeromonas hydrophila</italic> on biofilm formation

The RNA-binding protein Hfq is an evolutionarily conserved sRNA chaperone protein that can regulate the RNA translation at the post-transcription level and is involved in the regulation of important physiological functions in bacteria. The Aeromonas hydrophila ATCC 7966 genome contains two copies of the hfq gene ( AHA_0924 and AHA_3797 ). AHA_0924 has been found to share pronounced homology with many popular bacterial species and its biological functions was well documented. However, only few bacterial species carry the second hfq gene copy and its role remains largely unknown. To better understand the physiological function of AHA_3797 ( hfq2 ), we first successfully knocked out hfq2 using homologous recombination technology and found that its biofilm formation capability was significantly reduced in this study ( P ∆hfq2 in biofilm status. Our results showed a total of 1538 proteins, including 131 proteins that were up-regulated and 140 proteins that were down-regulated. Selected altered proteins were validated by Western blot analysis, which indicated the proteomics results were reliable. The following Gene Ontology (GO) annotation enrichment analysis in molecular functional clustering showed that up-regulated proteins were mainly involved in receptor activity, transaminase activity, and metal ion binding function, while down-regulated proteins were mainly involved in oxidoreductase activity. In the Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, the most plentiful up-regulated proteins tended to be involved in the biosynthesis of antibiotics, while the down-regulated proteins mainly involved chemotaxis of bacteria. We also found ∆hfq2 caused much of the up-regulation of bacterial iron transport-related proteins, such as colicin I receptor and TonB-dependent copper receptor, and down-regulation of chemotaxis-associated proteins, such as CheY, in the biofilm. The deletion of hfq2 did not result in a significant change in the expression of homologous Hfq protein, suggesting that the roles of both Hfq in the biofilm formation process of A. hydrophila are different. To verify the role of iron homeostasis during hfq2 -mediated biofilm formation, we compared the biofilm formation between ∆hfq2 and the wide-type strain under iron limitation conditions. The results showed that the biofilm formation of the control strain decreased, whereas that of ∆hfq2 i ncreased significantly after the addition of 200 μmol/L ferrous iron chelating 2,2′-dipyridyl (DIP), suggesting that Hfq2 protein negatively regulates intracellular iron homeostasis and affects biofilm formation. In general, this is the first work to confirm that the deletion of the hfq2 gene of A. hydrophila affects the formation of biofilm, suggesting that this gene is not a pseudogene, but rather it plays an important regulatory role in the formation of bacterial biofilm. Establishing the mechanism by which Hfq2 protein regulates bacterial biofilm formation would be of considerable scientific significance and may be helpful in the prevention and control of pathogenic bacteria and the development of new antibacterial strategies in the future.

Linoleic Acid Increases Prostaglandin E2 Release and Reduces Mitochondrial Respiration and Cell Viability in Human Trophoblast-Like Cells.

The omega 6 fatty acid (FA) linoleic acid (LA) is required for embryonic development; however, omega 6 FAs can alter cellular metabolism via inflammation or modulation of mitochondrial function. Fetal LA is obtained from the maternal diet, and FAs are transported to the fetus via placental FA transporters (FATPs) and binding proteins (FABPs), but specific proteins responsible for LA transport in placental trophoblasts are unknown. Dietary LA consumption is increasing, but the effect of elevated LA on trophoblast function is not clear. Swan71 trophoblasts were exposed to physiological and supraphysiological concentrations of LA for 24 hours. Quantification of mRNA was determined using real time PCR, and protein concentration was determined by Western blot analysis. Cell viability, citrate synthase activity and mitochondrial respiration were determined. Exposure to 300 and 500 μM LA increased FATP1 and FATP4 mRNA expression. 500 μM LA increased FATP1 and FATP4 protein expression. Exposure to 500 μM increased FABP5 mRNA expression, while exposure to 100 to 500 μM LA decreased FABP3 mRNA expression. 300 and 500 μM LA decreased FABP3 protein expression. Cell viability was decreased by exposure to LA (100 to 1000 μM). Citrate synthase activity and routine mitochondrial respiration were significantly decreased by exposure to 300 and 500 μM LA, and maximal respiration and spare respiratory capacity were decreased by exposure to 100 to 500 μM LA. 300 and 500 μM LA increased reactive oxygen species generation in human trophoblasts. Moreover, exposure to 300 and 500 μM LA decreased IL-6 secretion. Exposure to 500 μM LA increased IL-8, NF-κB and PPAR-γ mRNA expression, but decreased NF-κB protein expression. 300 μM LA decreased IL-8 protein expression. Further, exposure to 100 to 500 μM LA increased prostaglandin E2 and leukotriene B₄ release. Exposure to LA decreases cell viability, alters mRNA expression of FA transport related proteins, mitochondrial respiration and function, and inflammatory responses in trophoblasts. These findings may have implications on placental function when women consume high levels of LA.