Calmodulin-like protein CML15 interacts with PP2C46/65 to regulate papaya fruit ripening via integrating calcium, ABA and ethylene signals.

It is well known that calcium, ethylene and abscisic acid (ABA) can regulate fruit ripening, however, their interaction in the regulation of fruit ripening has not yet been fully clarified. The present study found that the expression of the papaya calcium sensor CpCML15 was strongly linked to fruit ripening. CpCML15 could bind Ca2+ and served as a true calcium sensor. CpCML15 interacted with CpPP2C46 and CpPP2C65, the candidate components of the ABA signalling pathways. CpPP2C46/65 expression was also related to fruit ripening and regulated by ethylene. CpCML15 was located in the nucleus and CpPP2C46/65 were located in both the nucleus and membrane. The interaction between CpCML15 and CpPP2C46/65 was calcium dependent and further repressed the activity of CpPP2C46/65 in vitro. The transient overexpression of CpCML15 and CpPP2C46/65 in papaya promoted fruit ripening and gene expression related to ripening. The reduced expression of CpCML15 and CpPP2C46/65 by virus-induced gene silencing delayed fruit colouring and softening and repressed the expression of genes related to ethylene signalling and softening. Moreover, ectopic overexpression of CpCML15 in tomato fruit also promoted fruit softening and ripening by increasing ethylene production and enhancing gene expression related to ripening. Additionally, CpPP2C46 interacted with CpABI5, and CpPP2C65 interacted with CpERF003-like, two transcriptional factors in ABA and ethylene signalling pathways that are closely related to fruit ripening. Taken together, our results showed that CpCML15 and CpPP2Cs positively regulated fruit ripening, and their interaction integrated the cross-talk of calcium, ABA and ethylene signals in fruit ripening through the CpCML15-CpPP2Cs-CpABI5/CpERF003-like pathway.

Novel mechanisms of cadmium tolerance and Cd-induced fungal stress in wheat: Transcriptomic and metagenomic insights.

Although several studies on the effects of cadmium (Cd) on wheat have been reported, the gene expression profiles of different wheat tissues in response to gradient concentrations of Cd, and whether soil microorganisms are involved in the damage to wheat remain to be discovered. To gain further insight into the molecular mechanisms of Cd-resistance in wheat, we sowed bread wheat (Triticum aestivum) in artificially Cd-contaminated soil and investigated the transcriptomic response of the wheat roots, stems, and leaves to gradient concentrations of Cd, as well as the alteration of the soil microbiome. Results indicated that the root bioaccumulation factors increased with Cd when concentrations were < 10 mg/kg, but at even higher concentrations, the bioaccumulation factors decreased, which is consistent with the overexpression of metal transporters and other genes related to Cd tolerance. In the Cd-contaminated soil, the abundance of fungal pathogens increased, and the antimicrobial response in wheat root was observed. Most of the differentially expressed genes (DEGs) of wheat changed significantly when the Cd concentration increased above 10 mg/kg, and the transcriptional response is much greater in roots than in stems and leaves. The DEGs are mainly involved in Cd transport and chelation, antioxidative stress, antimicrobial responses, and growth regulation. COPT3 and ZnT1 were identified for the first time as the major transporters responding to Cd in wheat. Overexpression of the nicotianamine synthase and pectinesterase genes suggested that nicotianamine and pectin are the key chelators in Cd detoxification. endochitinase, chitinase, and snakin2 were involved in the anti-fungal stress caused by Cd-induced cell damage. Several phytohormone-related DEGs are involved in the root's growth and repair. Overall, this study presents the novel Cd tolerance mechanisms in wheat and the changes in soil fungal pathogens that increase plant damage.

Metabolomic and Transcriptomic Profiling Provide Novel Insights into Fruit Ripening and Ripening Disorder Caused by 1-MCP Treatments in Papaya.

Treatment with 1-methylcyclopropylene (1-MCP) is an effective technique to preserve fruits, but inappropriate treatment with 1-MCP causes a ripening disorder (rubbery texture) in papaya fruit. In this study, a combined metabolomic and transcriptomic analysis was conducted to reveal the possible mechanism of the ripening disorder caused by unsuitable 1-MCP in papaya. A total of 203 differential accumulated metabolites (DAMs) were identified in the metabolome analysis. Only 24 DAMs were identified in the control (CK) vs. the 1-MCP 2 h group, and they were primarily flavonoids. Ninety and 89 DAMs were identified in the CK vs. 1-MCP 16 h and 1-MCP 2 h vs. 1-MCP 16 h groups, respectively, indicating that long-term 1-MCP treatment severely altered the metabolites during fruit ripening. 1-MCP 16 h treatment severely reduced the number of metabolites, which primarily consisted of flavonoids, lipids, phenolic acids, alkaloids, and organic acids. An integrated analysis of RNA-Seq and metabolomics showed that various energy metabolites for the tricarboxylic acid cycle were reduced by long-term treatment with 1-MCP, and the glycolic acid cycle was the most significantly affected, as well as the phenylpropane pathway. These results provide valuable information for fruit quality control and new insight into the ripening disorder caused by unsuitable treatment with 1-MCP in papaya.

The effect of trees on human energy fluxes in a humid subtropical climate region.

Trees are considered to be an effective tool for improving human thermal comfort in hot climates and have been widely used in landscape architecture. However, it is not always clear how trees affect human-environment energy fluxes. In this study, an in-depth analysis of four common tree species was undertaken based on comprehensive field measurements, in terms of how each tree and its characteristics affected the energy fluxes of a person in a humid subtropical climate region. Results showed that the largest effect of trees was on radiation fluxes, with a much smaller effect on the convective and evaporative fluxes. For a person standing in shade, a tree can reduce approximately 25% of the absorbed radiation compared with an open reference point. Moreover, the cooling effect on radiation components was found to be greater in the solar radiation domain than in the terrestrial radiation domain. Solar radiation and ground surface temperature had the largest effect on a human energy budget, which was affected by characteristics of the trees and the thermophysical properties of ground surfaces. The effect from relative humidity and wind speed was quite minimal. For the four common tree species in this study, Ficus microcarpa had the best thermal performance by reducing the most absorbed solar radiation flux. This study shows a detailed empirical research about the thermal effects of trees on a person, providing recommendations for tree species selection in urban design.

[Impact of PAHs on the expression of PRDX in earthworm (Eisenia fetida)].

A peroxiredoxin6 (PRDX6)-like gene segment has been found in up-regulated cDNA libraries of earthworm Eisenia fetida exposed to benzo[a]pyrene(BaP). Analysis with basic local alignment search tool (BLAST) and phylogenetic analysis revealed that the gene segment contains a characteristic motif which was encoded by Cys, illustrating that this gene segment belongs to PRDX. To verify the response of PRDX in E. feitida exposed to poly aromatic hydrocarbons (PAHs), four-week pollution experiments were conducted following the methods recommended by the Organization for Economic Co-operation and Development (OECD). The results showed that the expression of PRDX was up-regulated in earthworm exposed to 1.0 mg x kg(-1) Pyrene and BaP. It was indicated that PRDX in E. fetida was the potential molecular biomarkers of antioxidative stress caused by soil contamination.

Three novel cytochrome P450 genes identified in the marine polychaete Perinereis nuntia and their transcriptional response to xenobiotics.

Polychaetes have previously been used as bioindicators of environmental pollution. Their ability to eliminate organic pollutants such as polycyclic aromatic hydrocarbons (PAH) has been extensively analyzed. However, the cytochrome P450 monooxygenases (CYP) genes in polychaetes, which catalyze the first step of oxidative degradation of PAHs, have received little attention. Based on the partial sequences of three CYP genes that were enriched by subtractive cDNA libraries of the polychaete Perinereis nuntia, we amplified and sequenced the full-length cDNA of these novel CYP genes. These genes were named CYP4BB2, CYP423A1 and CYP424A1 by the Cytochrome P450 Nomenclature Committee. The deduced amino acid sequence of CYP4BB2 in P. nuntia showed 68% sequence identity to CYP4BB1 in Nereis virens, and was listed as a new member of the CYP4BB subfamily. The sequence of CYP423A1 and CYP424A1 both share less than 40% sequence identity to all known CYP enzymes and were classed into new CYP families. CYP family members are composite parts of a larger group called a clan. CYP4BB2 and CYP424A1 are listed as CYP4 clan members, while CYP423A1 is of the CYP2 clan. The 3D structures of these P. nuntia CYPs were successfully predicted by homology-modeling using the SWISS-MODEL workspace. The models of CYP424A1 and CYP4BB2 were created using 1jpzB (CYP102A) as a template, while CYP423A1 utilized 3czhB (CYP2R1) as its template. The presence of characteristic CYP superfamily motifs, such as the F-G⋯C-G amino acid sequence, and the conservation of the three-dimensional CYP structure shown by the modeling, suggested that these novel P. nuntia CYP genes may contain conserved functional domains of CYP monooxygenases. To examine the effect of xenobiotics on living organisms, we analyzed the transcriptional levels of these three new CYP genes in sandworms (P. nuntia) exposed to seawater artificially contaminated with benzo[a]pyrene (BaP). We also exposed individuals to industrial wastewater collected from Quanzhou Bay, Fujian, China, which was known to be contaminated with PAHs. Worms exposed to BaP had significantly higher levels of CYP4BB2, CYP423A1 and CYP424A1 mRNA. Transcription was up-regulated 5.9-, 5.3- and 12.3-folds respectively compared with the control worms living in clean seawater. The transcriptional levels of CYPs in worms cultured in the diluted wastewater collected from Quanzhou Bay, all positively correlated with the levels of PAHs detected in the water. The transcriptional up-regulation of the three CYP genes observed in this study, suggest the monooxygenases encoded by these CYP genes may play an important role in the detoxification of PAHs in this polychaete worm. These CYPs maybe essential for the adaptation of worms to contaminated environments and may be useful in the assessment of xenobiotic exposure.

[Composition and distribution of TCBS bacteria groups from sediments of Jiulong River estuary].

OBJECTIVE: To investigate potential pathogens in waters of Xiamen from Jiulong River, and to provide useful information for the prevention and control of potential pathogen infections. METHODS: All samples were spread on Thiosulfate Citrate Bile Salts Sucrose (TCBS) agar plates, and then incubated at 26 +/- 1 degrees C for 24 +/- 2 h. In total 158 TCBS strains were isolated from TCBS agar plates and pure-cultivated on 2216E agar plates. All strains were identified using the 16S rRNA gene- Restriction fragment length polymorphism (RFLP), 16S rRNA sequence analysis, GenBank database Basic Local Alignment Search Tool (BLAST) and phylogenetic analysis. RESULTS: The results show that 158 TCBS strains from the sediments of Jiulong River estuary were classfied as 7 genus, which were Pseudomonas (28%), Aeromonas (24%), Pseudoalteromonas (19%), Shewanella (13%), Bacillus (11%), Vibrio (4%) and Psychrobacter (1%). The composition and distribution of TCBS bacteria groups varied with stations. Non-halophilic or haloduric bacteria groups were dominant in the upper area of Jiulong River estuary, and halophilic and haloduric bacteria were dominant in the lower area, which characterized a typical estuary feature. The salinity played a key role in the distribution of TCBS groups. Vibrios did not constitute a significant proportion (6% - 19%) of the total TCBS strains at different stations, and most of the them distributed at the lower region. CONCLUSION: There were a lot of potential pathogens in Jiulong River estuary. Aeromonas, a typical genus of halotolerant bacteria, was the potentially terrigenous bacteria contamination to the waters of Xiamen. Most Vibrio specieses were marine aborigines, which was not directly contaminated from the runoff of Jiulong River.

Toxicity evaluation of benzo[a]pyrene on the polychaete Perinereis nuntia using subtractive cDNA libraries.

To gain insight into the toxic effects of the carcinogenic PAH benzo[a]pyrene (BaP) on the typical marine benthic polychaete Perinereis nuntia, we amplified and sequenced genes by creating subtractive cDNA libraries between worms exposed to BaP and solvent control. We assigned functions to the identified sequences and further analyzed the transcriptional profile changes of a set of 50 selected potential marker genes using quantitative real time PCR. A total of 2422 new high quality ESTs (GenBank accession number GT629654-GT632075) were obtained in the P. nuntia subtracted cDNA libraries, and assembled into 1594 unique sequences. Blastx results showed 700 of the unique sequences shared high similarity with existing genes in the GenBank nr database. Functional annotation of these enriched gene segments suggested that P. nuntia shows a wide range of toxicological responses to BaP. Comparison of the transcriptional profiles of the 50 potential marker genes in worms exposed to BaP and the control suggested that BaP significantly changed the expression of genes involved in xenobiotics metabolism, reactive oxygen species (ROS) elimination, DNA repair, apoptosis, cell division cycle, neurodegeneration, neurotransmitter metabolism and carcinogenesis. It also shows that there are significant correlations between these potential marker genes. The results support the prediction that the polychaete P. nuntia also has a set of tumor-related genes, while other responses influenced by BaP involve detoxification, antioxidation, DNA repair and apoptosis.

Antibiotics pollution in Jiulong River estuary: source, distribution and bacterial resistance.

To gain insight into the antibiotic pollution in the Jiulong River estuary and the pollutant sources, we analyzed the concentration of 22 widely-used antibiotics in water samples collected from the river and estuary, 17 and 18 sampling sites, respectively. Contamination with sulfonamides, quinolones and chloramphenicols was frequently detected and the distribution pattern of antibiotics suggested that most of the pollutants are from the Jiulong River, especially from the downstream watersheds. To reveal the ecological effects, we isolated 35 bacterial strains from the estuary and analyzed their antibiotic resistance to the eight most frequently detected antibiotics. The bacteria were subsequently classified into seven different genera by 16SrDNA sequencing. Up to 97.1% of the bacteria showed resistance and 70.6% of strains showed multi-resistance to these antibiotics, especially to sulfonamides. This study demonstrated a pattern of antibiotic contamination in the Jiulong River and its estuary and illustrated high bacterial antibiotic resistance which was significantly correlated with the average antibiotics concentrations and detected frequencies in the estuary.

Reproductive toxic effects of sublethal cadmium on the marine polychaete Perinereis nuntia.

To gain insight into the reproductive toxicity of sublethal cadmium on marine polychaetes, Perinereis nuntia sandworms were exposed to cadmium via artificially contaminated seawater. Cadmium influence on sexual maturation, egg laying, fertilization, zygote hatching and vitellogenin expression levels were analyzed. Results indicated that 23.05 and 563.87 microg L(-1) cadmium significantly delayed sexual maturation. Fertilization rate was significantly inhibited by 563.87 microg L(-1) cadmium while lower concentrations showed no significant effect. Zygote hatching was significantly inhibited by cadmium concentrations greater than 1.12 microg L(-1). We identified a vitellogenin gene sequence in P. nuntia and found that mRNA transcription was significantly upregulated by cadmium. These results indicate that sublethal cadmium levels cause dose-dependent reproductive toxicity on P. nuntia by inhibiting sexual maturation, fertilization and zygote hatching, and the increased expression of vitellogenin suggesting cadmium has strong feminization effects on polychaetes.

[Impact of benzo [a] pyrene the expression of mitochondrion-encoded genes in the earthworm Eisenia fetida].

The earthworm Eisenia fetida's benzo [a] pyrene (BaP) exposure experiments were carried out in artificial soil according to ISO 11268-1:1993. And then the upregulated and downregulated subtractive cDNA libraries were constructed by Clontech PCR-Select cDNA Subtration Kit. From the BaP exposure upregulated subtractive cDNA library, several cDNA segments matched mitochondrion-encoded genes were found, including cytochrome c oxidase subunit I (CO I), subunit II (CO II), subunit Ill (CO III), NADH dehydrogenase subunit 1 (NDH1), and ATP synthase subunit 6. The result indicated BaP and the subsequent oxidative stress disturbed the expression of mitochondrion-encoded genes, and this was potential biomarker for oxidative stress following xenobiotic exposure.

[Low dose benzo(a)pyrene up-regulated the transcription of HSP70 and HSP90 in Eisenia fetida].

To search for the molecular biomarkers of sub-lethal polycyclic aromatic hydrocarbons (PAHs)-contamination of soil, the subtractive cDNA libraries of earthworm Eisenia fetida exposed to benzo(a)pyrene (BaP) in artificial soil were constructed by suppression subtractive hybridization. After sequencing and analyzing with basic local alignment search tool (BLAST), two clones matching heat shock protein 70 k Da (HSP70) and one clone matching heat shock protein 90 k Da (HSP90) were isolated from the up-regulated library, and subsequently, the up-regulation of HSP70 and HSP90 was verified by real-time PCR in E. fetida exposed to 0.1 mg x kg(-1) and 1.0 mg x kg(-1) BaP. It was indicated that these two newly identified HSPs in E. fetida were the potential molecular biomarkers for soil contamination monitoring.

Annetocin and TCTP expressions in the earthworm Eisenia fetida exposed to PAHs in artificial soil.

The toxicity of sublethal polycyclic aromatic hydrocarbons (PAHs) levels in soils was assessed by testing their impact on expression of annetocin, a reproduction regulating gene, and translationally controlled tumor protein (TCTP), a tumorigenic response gene, in the earthworm Eisenia fetida cultured in artificial soil spiked with, phenanthrene (Phe), pyrene (Pyr), fluoranthene (Flu), or benzo(a)pyrene (Bap). Annetocin and TCTP were both up-regulated by 0.1 and 1.0 mg kg(-1) benzo(a)pyrene and TCTP was down-regulated by 10.0 mg kg(-1) phenanthrene. Weight loss and cocoon production of the worms were also analyzed. Only 10.0 mg kg(-1) phenanthrene impacted earthworm weight loss significantly and no significant differences on cocoon production were observed. Our study indicated that the potential ecotoxicity of sublethal PAHs in soil should not be neglected and mRNA transcription level in earthworms was a more sensitive indicator of PAHs exposure than traditional indexes using cocoon production as endpoints and/or using the whole-organism as the test materials.