Transcriptional Analysis of Pseudomonas stutzeri A1501 Associated with Host Rice

Pseudomonas stutzeriA1501, associative and endophytic nitrogen-fixing bacterium showed the capacity of colonization in the rice roots and considered as the good colonizer in the rice plant. The experiment was conducted to study the expression of genes potentiality relevant to the association of nitrogen fixing Pseudomonas stutzeri with host rice and reveal the molecular mechanism by which underlying interaction between bacteria and host rice. The bacteria were shown to be uniformly distributed on the rhizoplane of the root and the density of bacteria was found at the intercellular junction and micro colony developed on the surface of the epidermal cells and on the cellular junctions. Root exudates of rice were the major components of carbon and energy sources for bacteria. RT-PCR analyses of pilK, metE, rpoN and fdhE genes expression of P. stutzeri A1501 were performed at positive and negative (control) conditions. After 1 h, it was found that pilK, metE and rpoN transcription were increased 5.7, 6.4 and 3.4-fold, respectively, whereas in the fdhE gene has no expression. Consequently, after 4 h pilk, fdhE, metE and rpoN were decreased -1.9, -4.4, -0.2 and -0.8-fold, respectively. The gene pilK, expression was up-regulation after 1 h and down-regulation after 4 h that has twitching motility to convey the bacterial cell to point of attachment in to host plant. The gene expressions of the bacteria, pilK, metE, rpoN and fdhE were up- and down-regulated during the influence of root exudates which regulated the colonization of bacteria during plant-microbe interaction.

Transcriptome analysis of adherens junction pathway-related genes after peripheral nerve injury

The neural regeneration process is driven by a wide range of molecules and pathways. Adherens junctions are critical cellular junctions for the integrity of peripheral nerves. However, few studies have systematically characterized the transcript changes in the adherens junction pathway following injury. In this study, a rat model of sciatic nerve crush injury was established by forceps. Deep sequencing data were analyzed using comprehensive transcriptome analysis at 0, 1, 4, 7, and 14 days after injury. Results showed that most individual molecules in the adherens junctions were either upregulated or downregulated after nerve injury. The m RNA expression of ARPC1 B, ARPC3, TUBA8, TUBA1 C, CTNNA2, ACTN3, MET, HGF, NME1 and ARF6, which are involved in the adherens junction pathway and in remodeling of adherens junctions, was analyzed using quantitative real-time polymerase chain reaction. Most of these genes were upregulated in the sciatic nerve stump following peripheral nerve injury, except for CTNNA2, which was downregulated. Our findings reveal the dynamic changes of key molecules in adherens junctions and in remodeling of adherens junctions. These key genes provide a reference for the selection of clinical therapeutic targets for peripheral nerve injury.

Characterization and Pathogenicity of Pseudopestalotiopsis vietnamensis Causing Gray Blight of Wuyi Rock Tea(Camellia sinensis)in China and Specific Mechanisms of Disease Infection

Gray blight disease(GBD)causes significant losses in tea production in China.Although genes and biological processes involved in resistance to fungal disease in tea plants have been identified,specific mechanisms of the GBD infection process remain unknown.In this study,morphological and multi-gene(TEF-TUB-ITS)phylogenetic characteristics were used to identify isolate CLBB1 of Pseudopestalotiopsis vietnamensis.Pathogenicity tests confirmed that isolate CLBB1 from tea leaves caused GBD in the susceptible tea cultivar Wuyi Rock(Camellia sinensis var.sinensis cv.Shuixian).Spores began to germinate 24 h after infection(hai),and after 48 h,elongated fungal hyphae formed from a single conidium.Transcriptome analysis revealed that 482,517,and 369 genes were differentially expressed at 24,48,and 72 hai,respectively,in Wuyi Rock tea leaves.Isolate CLBB1 infection elicited phenotype-related responses and activated defense-related pathways,including plant–pathogen interaction,MAPK signaling,and plant hormone signal transduction,suggesting a possible mechanism underlying phenotype-based susceptibility to CLBB1.Thus,a new Ps.vietnamensis strain causing GBD in the tea cultivar‘Shuixian’was discovered in this study.Transcriptome analysis indicated that pathogen invasion activated chitin-related MAPK pathways and that tea plants required a hormone to restrict CLBB1.

Transcriptomics-based identification and characterization of genes related to sugar metabolism in‘Hongshuijing’pitaya

Sugar composition not only affects fruit flavor but is also an important determinant of fruit taste and consumer preference.In this study,changes in the sugar content and sugar-metabolizing enzymes were investigated from different sections of various fruit development phases of‘Hongshuijing’pitaya(Hylocereus monacanthus).Genes related to sugar metabolism were also screened by transcriptome analyses.The results indicated that glucose was the major sugar in mature pitaya fruit,and was mainly regulated by vacuolar acid invertase(VAI)and sucrose synthase(SS)(degradative direction).Sugar accumulation varied in pulp between different sections of the pitaya fruit.VAI,neutral invertase(NI)and SS(degradative direction)are crucial enzymes for sugar accumulation in pitaya.The expression of 17 genes related to sucrose metabolism obtained from seven databases[NCBI non-redundant protein database(Nr),NCBI non-redundant nucleotide sequence database(Nt),EuKaryotic Orthologous Groups(KOG),The Protein Families(Pfam),Kyoto Encyclopedia of Genes and Genomes(KEGG),Swiss-prot,and Gene Ontology(GO)]were analyzed in different pitaya pulp sections.HpVAI1 had the highest relative expression level on the 29th day after pollination(DAP).Positive correlations were found between HpVAI1 expression and VAI activity;HpNI4 and NI activity;HpSS2,HpSS5,and SS activity(synthetic direction),indicating that HpVAI1,HpNI4,and HpSS2 and HpSS5 were involved in the regulation of VAI,NI,and SS(synthetic direction),respectively.HpVAI1 and HpNI4 regulated sucrose degradation and the accumulation of glucose and fructose,while HpSS2 and HpSS5 regulated sucrose synthesis.These results suggest that HpVAI1 plays a key role in sugar metabolism during fruit development of‘Hongshuijing’pitaya.The results of this study provide new information about sugar metabolism in pitaya fruit that could help improve fruit quality and the breeding of new cultivars.

Expression and immunocytochemical analysis of Autographa californica nucleopolyhedrovirus （AcMNPV） orf74 gene

Autographa californica nucleopolyhedrovirus off74 （Ac74） is located between 62 311 and 63 108bp in the AcMNPV genome, which encodes 265 amino acid residues with a predicted 31 kDa molecular weight. The homologues of Ac74 were searched using BLASTP in protein databases, GenBank/EMBL and SWISS-PROT. The result revealed that deduced Ac74 protein was homologous to the predicted products from 10 lepidoptera NPV ORFs. The multiple sequence alignments of Ac74 and its 10 homologues manifested only one amino acid residue was completely conserved. The transcript analysis revealed that the transcript of Ac74 was detected from 24-72 hours post-infection （hpi）. The product of Ac74 was detected at 24 hpi and lasted until 72 hpi by Western blot using anti-Ac74 antiserum, consistent with reverse transcriptase polymerase chain reaction results. These results suggested Ac74 was expressed during the later stages of infection. The product of Ac74 was 31kDa in size, consistent with predicted molecular weight. The subcellular localization of Ac74 proteins manifested Ac74 protein in the cytoplasm, and was hardly present in the nucleus at 24 hpi. The fluorescence was also observed in polyhedra, except cytoplasm at 72 hpi. Together, Ac74 is a functional protein with 31kDa molecular weight and is located in the cytoplasm and the polyhedra.

Phenotype and metabolism alterations in PCB-degrading Rhodococcus biphenylivorans TG9^(T) under acid stress

Environmental acidification impairs microorganism diversity and their functions on substance transformation.Rhodococcus is a ubiquitously distributed genus for contaminant detoxification in the environment,and it can also adapt a certain range of pH.This work interpreted the acid responses from both phenotype and metabolism in strain Rhodococcus biphenylivorans TG9^(T)(TG9)induced at pH 3.The phenotype alterations were described with the number of culturable and viable cells,intracellular ATP concentrations,cell shape and entocyte,degradation efficiency of polychlorinated biphenyl(PCB)31 and biphenyl.The number of culturable cells maintained rather stable within the first 10 days,even though the other phenotypes had noticeable alterations,indicating that TG9 possesses certain capacities to survive under acid stress.The metabolism responses were interpreted based on transcription analyses with four treatments including log phase(LP),acid-induced(PER),early recovery after removing acid(RE)and later recovery(REL).With the overview on the expression regulations among the 4 treatments,the RE sample presented more upregulated and less downregulated genes,suggesting that its metabolism was somehow more active after recovering from acid stress.In addition,the response mechanism was interpreted on 10 individual metabolism pathways mainly covering protein modification,antioxidation,antipermeability,H+consumption,neutralization and extrusion.Furthermore,the transcription variations were verified with RT-qPCR on 8 genes with 24-hr,48-hr and 72-hr acid treatment.Taken together,TG9 possesses comprehensive metabolism strategies defending against acid stress.Consequently,a model was built to provide an integrate insight to understand the acid resistance/tolerance metabolisms in microorganisms.

Profiling biotoxicities of hexafluoropropylene oxide trimer acid with human embryonic stem cell-based assays

Hexafluoropropylene oxide trimer acid(HFPO-TA), an emerging replacement of perfluorooctanoic acid(PFOA), has recently been reported to be a potential environmental contaminant.Due to the similar structure to PFOA, HFPO-TA may cause comparable adverse effects on human health. Therefore, evaluating the toxic profiles of HFPO-TA has become an urgent task.In this study, we investigated the cytotoxicity and hepatoxicity of HFPO-TA using human embryonic stem cell(h ESC)-based assays. Results showed that HFPO-TA reduced h ESCs’ viability in a dose dependent manner, and the calculated IC50 for 24, 48 and 72 hr were 222.8,167.4, and 80.6 μmol/L, respectively. Significant intracellular ROS accumulation and mitochondrion membrane potential reduction were detected with HFPO-TA exposure, and increased apoptotic/necrotic cells were also observed in high dose of HFPO-TA treated group.Moreover, HFPO-TA at noncytotoxic concentrations also significantly impaired the functions of induced hepatocytes by diminishing cell glycogen storage ability and deregulating specific functional genes. Transcriptome sequencing analysis identified a set of hepatic associated biological processes responding to HFPO-TA exposure. PPAR was the most significantly enriched pathway. Genes including FGA, FGB, FGG, AHSG, HRG, ITIH2, ALB were characterized as hub genes by cyto Hubba plug-in. These data indicated that HFPO-TA is a potential hepatotoxicant, and may not be a safe replacement for PFOA.

Induced Deactivation of Genes Encoding Chlorophyll Biosynthesis Enzymes Disentangles Tetrapyrrole- Mediated Retrograde Signaling

In photosynthetic organisms, tetrapyrrole-mediated retrograde signals are proposed to contribute to a bal- anced nuclear gene expression （NGE） in response to metabolic activity in chloroplasts. We followed an experimental short- term approach that allowed the assessment of modified NGE during the first hours of specifically modified enzymatic steps of the Mg branch of tetrapyrrole biosynthesis, when pleiotropic effects of other signals can be avoided. In response to 24-h-induced silencing of CHLH, CHLM, and CHL27 encoding the CHLH subunit of Mg chelatase, the Mg protoporphyrin methyltransferase and Mg protoporphyrin monomethylester cyclase, respectively, deactivated gene expression rapidly led to reduced activity of the corresponding enzymes and altered Mg porphyrin levels. But NGE was not substantially altered. When these three genes were continuously inactivated for up to 4 d, changes of transcript levels of nuclear genes were determined. CHL27 silencing for more than 24h results in necrotic leaf lesions and modulated transcript levels of oxidative stress-responsive and photosynthesis-associated nuclear genes （PhANGs）. The prolonged deactivation of CHLH and CHLM results in slightly elevated transcript levels of PhANGs and tetrapyrrole-associated genes. These time-resolved studies indicate a complex scenario for the contribution of tetrapyrrole biosynthesis on NGE mediated by IO2-induced signaling and feedback-regulated ALA synthesis.