Organelle pH in the Arabidopsis Endomembrane System

The pH of intracellular compartments is essential for the viability of cells. Despite its relevance, little is known about the pH of these compartments. To measure pH in vivo, we have first generated two pH sensors by combining the improved-solubility feature of solubility-modified green fluorescent protein （GFP） （smGFP） with the pH-sensing capabil- ity of the pHluorins and codon optimized for expression in Arabidopsis. PEpHluorin （plant-solubility-modified ecliptic pHluorin） gradually loses fluorescence as pH is lowered with fluorescence vanishing at pH 6.2 and PRpHluorin （plant- solubility-modified ratiomatric pHluorin）, a dual-excitation sensor, allowing for precise measurements. Compartment- specific sensors were generated by further fusing specific sorting signals to PEpHluorin and PRpHluorin. Our results show that the pH of cytosol and nucleus is similar （pH 7.3 and 7.2）, while peroxisomes, mitochondrial matrix, and plastidial stroma have alkaline pH. Compartments of the secretory pathway reveal a gradual acidification, spanning from pH 7.1 in the endoplasmic reticulum （ER） to pH 5.2 in the vacuole. Surprisingly, pH in the trans-Golgi network （TGN） and mul- tivesicular body （MVB） is, with pH 6.3 and 6.2, quite similar. The inhibition of vacuolar-type H＋-ATPase （V-ATPase） with concanamycin A （ConcA） caused drastic increase in pH in TGN and vacuole. Overall, the PEpHluorin and PRpHluorin are excellent pH sensors for visualization and quantification of pH in vivo, respectively.     

Redox Regulation of Arabidopsis Mitochondrial Citrate Synthase

Citrate synthase has a key role in the tricarboxylic （TCA） cycle of mitochondria of all organisms, as it cata- lyzes the first committed step which is the fusion of a carbon-carbon bond between oxaloacetate and acetyl CoA. The regulation of TCA cycle function is especially important in plants, since mitochondrial activities have to be coordinated with photosynthesis. The posttranslational regulation of TCA cycle activity in plants is thus far almost entirely unexplored. Although several TCA cycle enzymes have been identified as thioredoxin targets in vitro, the existence of any thioredoxin-dependent regulation as known for the Calvin cycle, yet remains to be demonstrated. Here we have investigated the redox regulation of the Arabidopsis citrate synthase enzyme by site-directed mutagenesis of its six cysteine residues. Our results indicate that oxidation inhibits the enzyme activity by the formation of mixed disulfides, as the partially oxidized citrate synthase enzyme forms large redox-dependent aggregates. Furthermore, we were able to demonstrate that thioredoxin can cleave diverse intraas well as intermolecular disulfide bridges, which strongly enhances the activity of the enzyme. Activity measurements with the cysteine variants of the enzyme revealed important cysteine residues affecting total enzyme activity as well as the redox sensitivity of the enzyme.     

Strigolactone-Regulated Hypocotyl Elongation Is Dependent on Cryptochrome and Phytochrome Signaling Pathways in Arabidopsis

Seedling development including hypocotyl elongation is a critical phase in the plant life cycle. Light regula- tion of hypocotyl elongation is primarily mediated through the blue light photoreceptor cryptochrome and red/far-red light photoreceptor phytochrome signaling pathways, comprising regulators including COP1, HY5, and phytochrome- interacting factors （PIFs）. The novel phytohormones, strigolactones, also participate in regulating hypocotyl growth. However, how strigolactone coordinates with light and photoreceptors in the regulation of hypocotyl elongation is largely unclear. Here, we demonstrate that strigolactone inhibition of hypocotyl elongation is dependent on cryp- tochrome and phytochrome signaling pathways. The photoreceptor mutants cry1 cry2, phyA, and phyB are hyposensi- tive to strigolactone analog GR24 under the respective monochromatic light conditions, while cop1 and pifl pif3 pif4 pif5 （pifq） quadruple mutants are hypersensitive to GR24 in darkness. Genetic studies indicate that the enhanced respon- siveness of cop1 to GR24 is dependent on HY5 and MAX2, while that of pifq is independent of HY5. Further studies demonstrate that GR24 constitutively up-regulates HY5 expression in the dark and light, whereas GR24-promoted HY5 protein accumulation is light- and cryptochrome and phytochrome photoreceptor-dependent. These results suggest that the light dependency of strigolactone regulation of hypocotyl elongation is likely mediated through MAX2-dependent promotion of HY5 expression, light-dependent accumulation of HY5, and PIF-regulated components.     

Cybertaxonomy to accomplish big things in aphid systematics

Biodiversity sciences have progressed at such a pace that the taxonomic community has been unable to grow concomitantly to keep up with the influx of biologicaldata. This ＂taxonomic impediment＂ has led some to suggest that taxonomy is no longer pertinent and to the development of methodologies that circumvent the taxonomic process.This article does not seek to argue for the importance of taxonomy but rather is a call to the aphid taxonomy community to rise to the challenge by dramatically increasing the volume and comprehensiveness of its output without sacrificing quality. Recent informatics technology allows us to mobilize the 2 most important aphid taxonomy resources： expertsand specimens, both distributed globally. ＂Cyberspecimens,＂ museum specimens digitally rendered at a resolution sufficient for remote identification, and open ＂cybertaxonomic＂tools will allow the international aphid taxonomic community to carry out large, ambitious, projects. The global aphid cybertaxonomy proposed here will serve not only the ends ofresearch aphidologists, but also provide a model for other taxonomic communities to adapt and adopt as we confront both the taxonomic impediment and the taxonomic naysayers.     

Plastid Signals and the Bundle Sheath： Mesophyll Development in Reticulate Mutants

The development of a plant leaf is a meticulously orchestrated sequence of events producing a complex organ comprising diverse cell types. The reticulate class of leaf variegation mutants displays contrasting pigmentation between veins and interveinal regions due to specific aberrations in the development of mesophyll cells. Thus, the reticulate mutants offer a potent tool to investigate cell-type-specific developmental processes. The discovery that most mutants are affected in plastid-localized, metabolic pathways that are strongly expressed in vasculature-associated tis- sues implicates a crucial role for the bundle sheath and their chloroplasts in proper development of the mesophyll cells. Here, we review the reticulate mutants and their phenotypic characteristics, with a focus on those in Arabidopsis thali- ana. Two alternative models have been put forward to explain the relationship between plastid metabolism and meso- phyll cell development, which we call here the supply and the signaling hypotheses. We critically assess these proposed models and discuss their implications for leaf development and bundle sheath function in C3 species. The characteriza- tion of the reticulate mutants supports the significance of plastid retrograde signaling in cell development and highlights the significance of the bundle sheath in C3 photosynthesis.     

Emerging roles of miR-210 and other non-coding RNAs in the hypoxic response

Hypoxia is a key component of the tumor microenviron- merit and represents a well-documented source of thera- peutic failure in clinical oncology. Recent work has provided support for the idea that non-coding RNAs, and in particular, microRNAs, may play important roles in the adaptive response to low oxygen in tumors. Specifically, all published studies agree that the induction of microRNA- 210 （miR-210） is a consistent feature of the hypoxic re- sponse in both normal and malignant cells, miR-210 is a robust target of hypoxia-inducible factors, and its overex- pression has been detected in a variety of diseases with a hypoxic component, including most solid tumors. High levels of miR-210 have been linked to an in vivo hypoxic sig- nature and to adverse prognosis in breast and pancreatic cancer patients. A wide variety of miR-210 targets have been identified, pointing to roles in mitochondrial metabol- ism, angiogenesis, DNA damage response, apoptosis, and cell survival. Such targets are suspected to affect the devel- opment of tumors in multiple ways; therefore, an increased knowledge about miR-210＇s functions may lead to novel diagnostic and therapeutic approaches in cancer.     

Dissecting the Heat Stress Response in Chlamydomonas by Pharmaceutical and RNAi Approaches Reveals Conserved and Novel Aspects

To study how conserved fundamental concepts of the heat stress response （HSR） are in photosynthetic eukaryotes, we applied pharmaceutical and antisense/amiRNA approaches to the unicellular green alga Chlamydomonas reinhardtii. The Chlamydomonas HSR appears to be triggered by the accumulation of unfolded proteins, as it was induced at ambient temperatures by feeding cells with the arginine analog canavanine. The protein kinase inhibitor staurosporine strongly retarded the HSR, demonstrating the importance of phosphorylation during activation of the HSR also in Chlamydomonas. While the removal of extracellular calcium by the application of EGTA and BAPTA inhibited the HSR in moss and higher plants, only the addition of BAPTA, but not of EGTA, retarded the HSR and impaired thermotoler- ance in Chlamydomonas. The addition of cycloheximide, an inhibitor of cytosolic protein synthesis, abolished the attenu- ation of the HSR, indicating that protein synthesis is necessary to restore proteostasis. HSP90 inhibitors induced a stress response when added at ambient conditions and retarded attenuation of the HSR at elevated temperatures. In addition, we detected a direct physical interaction between cytosolic HSP90A/HSP70A and heat shock factor 1, but surprisingly this interaction persisted after the onset of stress. Finally, the expression of antisense constructs targeting chloroplast HSP70B resulted in a delay of the cell＇s entire HSR, thus suggesting the existence of a retrograde stress signaling cascade that is desensitized in HSP7OB-antisense strains.     

Aphis （Hemiptera： Aphididae） species groups found in the Midwestern United States and their contribution to the phylogenetic knowledge of the genus

A phylogeny of the genus Aphis Linnaeus, 1 758 was built primarily from specimens collected in the Midwest of the United States. A data matrix was constructedwith 68 species and 41 morphological characters with respective character states of alate and apterous viviparous females. Dendrogram topologies of analyses performed usingUPGMA （Unweighted Pair Group Method with Arithmetic Mean）, Maximum Parsimony and Bayesian analysis of Cytochrome Oxidase I, Elongation Factor 1-α and primary endosymbiont Buchnera aphidicola 16S sequences were not congruent. Bayesian analysis strongly supported most terminal nodes of the phylogenetic trees. The phylogeny wasstrongly supported by EFI-α, and analysis of COl and EFI-α molecular data combined with morphological characters. It was not supported by single analysis of COI or Buch-hera aphidicola 16S. Results from the Bayesian phylogeny show 4 main species groups： asclepiadis,fabae, gossypii, and middletonii. Results place Aphis and species of the generaProtaphis Bōrner, 1952, Toxoptera Koch, 1856 and Xerobion Nevsky, 1928 in a monophyletic clade. Morphological characters support this monophyly as well. The phylogenyshows that the monophyletic clade of the North American middletonii species group belong to the genus Protaphis： P. debilicornis （Gillette ＆ Palmer, 1929）, comb. nov., P. echinaceae（Lagos and Voegtlin, 2009）, comb. nov., and P. middletonii （Thomas, 1879）. The genus Toxoptera should be considered a subgenus of Aphis （stat. nov.）. The analysis also indicatesthat the current genus Iowana Frison, 1954 should be considered a subgenus of Aphis （stat. nov.）.     

Cell Wall,Cytoskeleton, and Cell Expansion in Higher Plants

To accommodate two seemingly contradictory biological roles in plant physiology, providing both the rigid structural support of plant cells and the adjustable elasticity needed for cell expansion, the composition of the plant cell wall has evolved to become an intricate network of cellulosic, hemicellulosic, and pectic polysaccharides and protein. Due to its complexity, many aspects of the cell wall influence plant cell expansion, and many new and insightful observations and technologies are forthcoming. The biosynthesis of cell wall polymers and the roles of the variety of proteins involved in polysaccharide synthesis continue to be characterized. The interactions within the cell wall polymer network and the modification of these interactions provide insight into how the plant cell wall provides its dual function. The complex cell wall architecture is controlled and organized in part by the dynamic intracellular cytoskeleton and by diverse trafficking pathways of the cell wall polymers and cell wall-related machinery. Meanwhile, the cell wall is continually influenced by hormonal and integrity sensing stimuli that are perceived by the cell. These many processes cooperate to construct, maintain, and manipulate the intricate plant cell wall--an essential structure for the sustaining of the plant stature, growth, and life.     

How does the host-specialized aphid deal with food deficiency？

Aphis gossypii Glover shows obvious host specialization, with cucurbit- and cotton-specialized biotypes or host races in many regions. Because its annual natal hostcrops senesce earlier the cucurbit-specialized biotype may suffer food deficiency. The method this biotype uses to overcome this challenge is still poorly understood. In orderto understand the potential of the cucurbit-specialized biotype aphids in host shift and usage, the performance of this biotype on cotton （Gossypium hirsutum）, a common butpoor quality host plant, was explored in this study. The cucurbit-specialized aphids could establish populations on cotton only when these plants had at least nine leaves, and subsequent populations developed rather slowly. The presence of whitefly populations on cotton improved the success rate of cucurbit-specialized aphids. The cucurbit-specialized aphidswere mainly distributed on the older leaves of cotton, with only a few settling on the upper leaves. The cucurbit-specialized aphids reared on cotton for 40, 54 and 61 days stillmaintained strong preference for their natal host plant, cucumber （Cucumis sativus）, rather than cotton, and their net reproductive rates and intrinsic rates of natural increase weredramatically lower when they were transferred onto new six-leaf cotton plants or detached leaves. Therefore, we concluded that the cucurbit-specialized aphids have the potentialto utilize mature or whitefly-stressed cotton plants, but that this feeding experience on cotton did not alter their specialization for cucurbits. Some cotton plants could act as atemporary host for the cucurbit-specialized aphids to overcome food deficiency arising from senescing cucurbits.     

Regulation of Cytokinesis by Exocyst Subunit SEC6 and KEULE in Arabidopsis thaliana

Proper vesicle tethering and membrane fusion at the cell plate are essential for cytokinesis. Both the vesicle tethering complex exocyst and membrane fusion regulator KEULE were shown to function in cell plate formation, but the exact mechanisms still remain to be explored. In this study, using yeast two-hybrid （Y-2-H） assay, we found that SEC6 interacted with KEULE, and that a small portion of C-terminal region of KEULE was required for the interaction. The direct SEC6-KEULE interaction was supported by further studies using in vitro pull-down assay, immunoprecipitation, and in vivo bimolecular florescence complementation （BIFC） microscopy, sec6 mutants were male gametophytic lethal as reported; however, pollen-rescued sec6 mutants （PRsec6） displayed cytokinesis defects in the embryonic cells and later in the leaf pavement cells and the guard cells. SEC6 and KEULE proteins were co-localized to the cell plate during cytokine- sis in transgenic Arabidopsis. Furthermore, only SEC6 but not other exocyst subunits located in the cell plate interacted with KEULE in vitro. These results demonstrated that, like KEULE, SEC6 plays a physiological role in cytokinesis, and the SEC6-KEULE interaction may serve as a novel molecular linkage between arriving vesicles and membrane fusion machin- ery or directly regulate membrane fusion during cell plate formation in plants.     

Perturbation of Auxin Homeostasis Caused by Mitochondrial FtSH4 Gene-Mediated Peroxidase Accumulation Regulates Arabiclopsis Architecture

Reactive oxygen species and auxin play important roles in the networks that regulate plant development and morphogenetic changes, However, the molecular mechanisms underlying the interactions between them are poorly understood. This study isolated a mas （More Axillary Shoots） mutant, which was identified as an allele of the mitochondrial AAA-protease AtFtSH4, and characterized the function of the FtSH4 gene in regulating plant development by medi- ating the peroxidase-dependent interplay between hydrogen peroxide （H2Oz） and auxin homeostasis. The phenotypes of dwarfism and increased axillary branches observed in the mas （renamed as ftsh4-4） mutant result from a decrease in the IAA concentration. The expression levels of several auxin signaling genes, including IAA1, IAA2, and IAA3, as well as several auxin binding and transport genes, decreased significantly in ftsh4-4 plants. However, the H202 and peroxidases levels, which also have IAA oxidase activity, were significantly elevated in ftsh4-4 plants. The ftsh4-4 phenotypes could be reversed by expressing the iaaM gene or by knocking down the peroxidase genes PRX34 and PRX33. Both approaches can increase auxin levels in the ftsh4-4 mutant. Taken together, these results provided direct molecular and genetic evidence for the interaction between mitochondrial ATP-dependent protease, H2O2, and auxin homeostasis to regulate plant growth and development.     

A New Set of Reversibly Photoswitchable Fluorescent Proteins for Use in Transgenic Plants

Fluorescent reporter proteins that allow repeated switching between a fluorescent and a non-fluorescent state in response to specific wavelengths of light are novel tools for monitoring of protein trafficking and super-resolu- tion fluorescence microscopy in living organisms. Here, we describe variants of the reversibly photoswitchable fluores- cent proteins rsFastLime, bsDronpa, and Padron that have been codon-optimized for the use in transgenic Arabidopsis plants. The synthetic proteins, designated rsFastLIME-s, bsDRONPA-s, and PADRON C-s, showed photophysical properties and switching behavior comparable to those reported for the original proteins. By combining the ＇positively switchable＇ PADRON C-s with the ＇negatively switchable＇ rsFastLIME-s or bsDRONPA-s, two different fluorescent reporter proteins could be imaged at the same wavelength upon transient expression in Nicotiana benthamiana cells. Thus, co-localiza- tion analysis can be performed using only a single detection channel. Furthermore, the proteins were used to tag the RNA-binding protein AtGRP7 （Arabidopsis thaliana glycine-rich RNA-binding protein 7） in transgenic Arabidopsis plants. Because the new reversibly photoswitchable fluorescent proteins show an increase in signal strength during each pho- toactivation cycle, we were able to generate a large number of scans of the same region and reconstruct 3-D images of AtGRP7 expression in the root tip. Upon photoactivation of the AtGRP7：rsFastLIME-s fusion protein in a defined region of a transgenic Arabidopsis root, spreading of the fluorescence signal into adjacent regions was observed, indicating that movement from cell to cell can be monitored. Our results demonstrate that rsFastLIME-s, bsDRONPA-s, and PADRON C-s are versatile fluorescent markers in plants, Furthermore, the proteins also show strong fluorescence in mammalian cells including COS-7 and HeLa cells.     

A trait-based approach to assess climate change sensitivity of freshwater invertebrates across Swedish ecoregions

Freshwater habitats and organisms are among the most threatened on Earth, and freshwater ecosystems have been subject to large biodiversity losses. We developed a Climate Change Sensitivity （CCS） indicator based on trait information for a selection of stream- and lake-dwelling Ephemeroptera, Plecoptera and Trichoptera taxa. We calculated the CCS scores based on ten species traits identified as sensitive to global climate change. We then assessed climate change sensitivity between the six main ecoregions of Sweden as well as the three Swedish regions based on lilies. This was done using biological data from 1,382 stream and lake sites where we compared large-scale （ecoregional） patterns in climate change sensitivity with potential future exposure of these ecosystems to increased temperatures using ensemble-modelled future changes in air temperature. Current （1961-1990） measured temperature and ensemble-modelled future （2100） temperature showed an increase from the northernmost towards the southern ecoregions, whereas the predicted temperature change increased from south to north. The CCS indicator scores were highest in the two northernmost boreal ecoregions where we also can expect the largest global climate change-induced increase in temperature, indicating an unfortunate congruence of exposure and sensitivity to climate change. These results are of vital importance when planning and implementing management and conservation strategies in freshwater ecosystems, e.g., to mitigate increased temperatures using riparian buffer strips. We conclude that traits information on taxa specialization, e.g., in terms of feeding specialism or taxa having a preference for high altitudes as well as sensitivity to changes in temperature are important when assessing the risk from future global climate change to freshwater ecosystems [Current Zoology 60 （2）： 221-232, 2014].     

A New β-Estradiol-lnducible Vector Set that Facilitates Easy Construction and Efficient Expression of Transgenes Reveals CBL3-Dependent Cytoplasm to Tonoplast Translocation of CIPK5

Transient and stable expression of transgenes is central to many investigations in plant biology research. Chemical regulation of expression can circumvent problems of plant lethality caused by constitutive overexpression or allow inducible knock （out/down） approaches. Several chemically inducible or repressible systems have been described and successfully applied. However, cloning and application-specific modification of most available inducible expression systems have been limited and remained complicated due to restricted cloning options. Here we describe a new set of 57 vectors that enable transgene expression in transiently or stably transformed cells. All vectors harbor a synthetically optimized XVE expression cassette, allowing I~-estradiol mediated protein expression. Plasmids are equipped with the reporter genes GUS, GFP, mCherry, or with HA and Strepll epitope tags and harbor an optimized multiple cloning site for flexible and simple clon- ing strategies. Moreover, the vector design allows simple substitution of the driving promoter to achieve tissue-specificity or to modulate expression ranges of inducible transgene expression. We report details of the kinetics and dose-dependence of expression induction in Arabidopsis leaf mesophyll protoplasts, transiently transformed Nicotiana benthamiana leaves, and stably transformed Arabidopsis plants. Using these vectors, we investigated the influence of CBL （Calcineurin B-like） protein expression on the subcellular localization of CIPKs （Calcineurin B-like interacting protein kinases）. These analyses uncovered that induced co-expression of CBL3 is fully sufficient for dynamic translocation of CIPK5 from the cytoplasm to the tonoplast. Thus, the vector system presented here facilitates a broad range of research applications.     

Heteromeric and Homomeric Geranyl Diphosphate Synthases from Catharanthus roseus and Their Role in Monoterpene Indole Alkaloid Biosynthesis

Catharanthus roseus is the sole source of two most important monoterpene indole alkaloid （MIA） anti- cancer agents： vinblastine and vincristine. MIAs possess a terpene and an indole moiety derived from terpenoid and shikimate pathways, respectively. Geranyl diphosphate （GPP）, the entry point to the formation of terpene moiety, is a product of the condensation of isopentenyl diphosphate （IPP） and dimethylallyl diphosphate （DMAPP） by GPP synthase （GPPS）. Here, we report three genes encoding proteins with sequence similarity to large subunit （CrGPPS.LSU） and small subunit （CrGPPS.SSU） of heteromeric GPPSs, and a homomeric GPPSs. CrGPPS.LSU is a bifunctional enzyme producing both GPP and geranyl geranyl diphosphate （GGPP）, CrGPPS.SSU is inactive, whereas CrGPPS is a homomeric enzyme forming GPP. Co-expression of both subunits in Escherichia coil resulted in heteromeric enzyme with enhanced activity producing only GPR While CrGPPS.LSU and CrGPPS showed higher expression in older and younger leaves, respectively, CrGPPS.SSU showed an increasing trend and decreased gradually. Methyl jasmonate （MelA） treatment of leaves sig- nificantly induced the expression of only CrGPPS.SSU. GFP localization indicated that CrGPPS.SSU is plastidial whereas CrGPPS is mitochondrial. Transient overexpression of AmGPPS.SSU in C. roseus leaves resulted in increased vindoline, immediate monomeric precursor of vinblastine and vincristine. Although C. roseus has both heteromeric and homomeric GPPS enzymes, our results implicate the involvement of only heteromeric GPPS with CrGPPS.SSU regulating the GPP flux for MIA biosynthesis.     

ERK1/2 mediates lung adenocarcinoma cell proliferation and autophagy induced by apelin-13

The aim of this study was to investigate the role of apelin in the cell proliferation and autophagy of lung adenocarcin- oma. The over-expression of APJ in lung adenocarcinoma was detected by immunohistochemistry, while plasma apelin level in lung cancer patients was measured by enzyme-linked immunosorbent assay. Our findings revealed that apelin-13 significantly increased the phosphorylation of ERK1/2, the expression of cyclin D1, microtubule-associated protein 1 light chain 3A/B （LC3A/B）, and beclinl, and con- fwmed that apelin-13 promoted A549 cell proliferation and induced A549 cell autophagy via ERK1/2 signaling. More- over, there are pores on the surface of human lung adeno- carcinoma cell line A549 and apelin-13 causes cell surface smooth and glossy as observed under atomic force micros- copy. These results suggested that ERK1/2 signaling pathway mediates apelin-13-induced lung adenocarcinoma cell proliferation and autophagy. Under our experimental condition, autophagy associated with 3-methyladenine was not involved in cell proliferation.     

Arabidopsis Coordinates the Diurnal Regulation of Carbon Allocation and Growth across a Wide Range of Photoperiods

In short photoperiods, plants accumulate starch more rapidly in the light and degrade it more slowly at night, ensuring that their starch reserves last until dawn. To investigate the accompanying changes in the timing of growth, Arabidopsis was grown in a range of photoperiods and analyzed for rosette biomass, photosynthesis, respiration, ribosome abundance, polysome loading, starch, and over 40 metabolites at dawn and dusk. The data set was used to model growth rates in the daytime and night, and to identify metabolites that correlate with growth. Modeled growth rates and polysome loading were high in the daytime and at night in long photoperiods, but decreased at night in short photoperiods. Ribosome abundance was similar in all photoperiods. It is discussed how the amount of starch accumulated in the light period, the length of the night, and maintenance costs interact to constrain growth at night in short photoperiods, and alter the strategy for optimizing ribosome use. Significant correlations were found in the day- time and the night between growth rates and the levels of the sugar-signal trehalose 6-phosphate and the amino acid biosynthesis intermediate shikimate, identifying these metabolites as hubs in a network that coordinates growth with diurnal changes in the carbon supply.     

Genotoxicity biomarkers in aquatic bioindicators

Pollution of the aquatic environment is an ever-growing problem, as waters are the ultimate sink for the large number of xenobiotics from multiple sources. DNA damaging agents have a significant ecological relevance since they are implicated in many pathological processes and exert effects beyond that of individual being active through following generations. A large number of methods have been applied to evaluate genotoxic damage in different aquatic species. Comet assay, as method for de- tecting DNA alterations, and micronucleus test, as an index of chromosomal damage are the most widely applied and validated methods in field studies. These methods were applied in different vertebrate and invertebrate aquatic species, but only mollusk and fish species have been employed in routine biomonitoring programs. Mussels, due to their widely geographical distribution and the suitability for caging represent the bioindicator of choice in field studies. Mytilus species is the most used marine mussel. The use of fish is limited to specific geographic areas. The present review mainly focuses on the application of comet assay and micronucleus test in mussels. A number of biomonitoring studies in mussels, using comet assay or micronucleus test, revealed exposure to different classes of genotoxic compounds with a good discrimination power. The different evidence from the two as- says, reflects different biological mechanisms for the two genetic endpoints, DNA damage and chromosomal damage, suggesting their combined application in the field. Different endogenous and exogenous factors have been shown to modulate the genotoxic responses in mussels, acting as confounding factors in environmental monitoring. The use of standardized protocol for caging, sampling and genotoxity evaluation is critical in biomonitoring studies. The use of a multimarker approach coupling genotoxicity biomarkers with physiological and biochemical factors allows to have a complete picture of the environmental pollution [Current Zoology 60 （2）： 273-284, 2014].