Effect of Particle Wettability and Particle Concentration on the Enzymatic Dehydration of n-Octanaloxime in Pickering Emulsions.

Pickering emulsion systems have emerged as platforms for the synthesis of organic molecules in biphasic biocatalysis. Herein, the catalytic performance was evaluated for biotransformation using whole cells exemplified for the dehydration of n-octanaloxime to n-octanenitrile catalysed by an aldoxime dehydratase (OxdB) overexpressed in E. coli. This study was carried out in Pickering emulsions stabilised solely with silica particles of different hydrophobicity. We correlate, for the first time, the properties of the emulsions with the conversion of the reaction, thus gaining an insight into the impact of the particle wettability and particle concentration. When comparing two emulsions of different type with similar stability and droplet diameter, the oil-in-water (o/w) system displayed a higher conversion than the water-in-oil (w/o) system, despite the conversion in both cases being higher than that in a "classic" two-phase system. Furthermore, an increase in particle concentration prior to emulsification resulted in an increase of the interfacial area and hence a higher conversion.

Identification and Functional Analysis of Tomato CIPK Gene Family.

The calcineurin B-like interacting protein kinase (CIPK) protein family is a critical protein family in plant signaling pathways mediated by Ca2+, playing a pivotal role in plant stress response and growth. However, to the best of our knowledge, no study of the tomato CIPK gene family in response to abiotic stress has been reported. In this study, 22 members of the tomato CIPK gene family were successfully identified by using a combination of bioinformatics techniques and molecular analyses. The expression level of each member of tomato CIPK gene family under abiotic stress (low temperature, high salt, drought treatment) was determined by qRT-PCR. Results indicated that tomato CIPK demonstrated different degrees of responding to various abiotic stresses, and changes in SlCIPK1 and SlCIPK8 expression level were relatively apparent. The results of qRT-PCR showed that expression levels of SlCIPK1 increased significantly in early stages of cold stress, and the expression level of SlCIPK8 increased significantly during the three treatments at different time points, implicating Solanum lycopersicum CIPK1(SlCIPK1) and Solanum lycopersicum CIPK8 (SlCIPK8) involvement in abiotic stress response. SlCIPK1 and SlCIPK8 were silenced using Virus-induced gene silencing (VIGS), and physiological indexes were detected by low temperature, drought, and high salt treatment. The results showed that plants silenced by SlCIPK1 and SlCIPK8 at the later stage of cold stress were significantly less resistant to cold than wild-type plants. SlCIPK1 and SlCIPK8 silenced plants had poor drought resistance, indicating a relationship between SlCIPK1 and SlCIPK8 with response to low temperature and drought resistance. This is the first study to uncover the nucleotide sequence for tomato CIPK family members and systematically study the changes of tomato CIPK family members under abiotic stress. Here, we investigate the CIPK family's response under abiotic stress providing understanding into the signal transduction pathway. This study provides a theoretical basis for elucidating the function of tomato CIPK at low temperature and its molecular mechanism of regulating low temperatures.

Insight into transketolase of Pyropia haitanensis under desiccation stress based on integrative analysis of omics and transformation.

BACKGROUND: Pyropia haitanensis, distributes in the intertidal zone, can tolerate water losses exceeding 90%. However, the mechanisms enabling P. haitanensis to survive harsh conditions remain uncharacterized. To elucidate the mechanism underlying P. haitanensis desiccation tolerance, we completed an integrated analysis of its transcriptome and proteome as well as transgenic Chlamydomonas reinhardtii carrying a P. haitanensis gene. RESULTS: P. haitanensis rapidly adjusted its physiological activities to compensate for water losses up to 60%, after which, photosynthesis, antioxidant systems, chaperones, and cytoskeleton were activated to response to severe desiccation stress. The integrative analysis suggested that transketolase (TKL) was affected by all desiccation treatments. Transgenic C. reinhardtii cells overexpressed PhTKL grew better than the wild-type cells in response to osmotic stress. CONCLUSION: P. haitanensis quickly establishes acclimatory homeostasis regarding its transcriptome and proteome to ensure its thalli can recover after being rehydrated. Additionally, PhTKL is vital for P. haitanensis desiccation tolerance. The present data may provide new insights for the breeding of algae and plants exhibiting enhanced desiccation tolerance.

The contribution of collecting duct NOS1 to the concentrating mechanisms in male and female mice.

The collecting duct (CD) concentrates the urine, thereby maintaining body water volume and plasma osmolality within a normal range. The endocrine hormone arginine vasopressin acts in the CD to increase water permeability via the vasopressin 2 receptor (V2R)-aquaporin (AQP) axis. Recent studies have suggested that autocrine factors may also contribute to the regulation of CD water permeability. Nitric oxide is produced predominantly by nitric oxide synthase 1 (NOS1) in the CD and acts as a diuretic during salt loading. The present study sought to determine whether CD NOS1 regulates diuresis during changes in hydration status. Male and female control and CD NOS1 knockout (CDNOS1KO) mice were hydrated (5% sucrose water), water deprived, or acutely challenged with the V2R agonist desmopressin. In male mice, water deprivation resulted in decreased urine flow and increased plasma osmolality, copeptin concentration, and kidney AQP2 abundance independent of CD NOS1. In female control mice, water deprivation reduced urine flow, increased plasma osmolality and copeptin, but did not significantly change total AQP2; however, there was increased basolateral AQP3 localization. Surprisingly, female CDNOS1KO mice while on the sucrose water presented with symptoms of dehydration. Fibroblast growth factor 21, an endocrine regulator of sweetness preference, was significantly higher in female CDNOS1KO mice, suggesting that this was reducing their drive to drink the sucrose water. With acute desmopressin challenge, female CDNOS1KO mice failed to appropriately concentrate their urine, resulting in higher plasma osmolality than controls. In conclusion, CD NOS1 plays only a minor role in urine-concentrating mechanisms.

Effects of dehydration on echocardiographic diastolic parameters in healthy cats.

This study aimed to assess the effects of dehydration on echocardiographic indices in healthy cats: specifically, it aimed to assess the effects of volume depletion on diastolic function. Nine experimental cats were subjected to both a dehydration and placebo protocol separated by a 21-day washout period. Echocardiography was performed at baseline and on completion of each protocol. Results were compared between the two protocols. Volume depletion was induced by intravenous administration of furosemide. Volume depletion showed a significant association with increased interventricular septal and left ventricular free wall thickness at end-diastole, decreased left ventricular internal diameter at end-diastole, and left atrial diameter at end-systole. The peak early (E) and late (A) diastolic filling velocities, and the peak early diastolic velocities (E') were significantly decreased by dehydration. Volume depletion did not affect peak longitudinal strain rate during early diastole, E/A, or E/E'. Volume depletion significantly affected the echocardiographic diastolic indices and conventional echocardiographic parameters in healthy cats.

Functional characterization of galactinol synthase and raffinose synthase in desiccation tolerance acquisition in developing Arabidopsis seeds.

Raffinose family oligosaccharides (RFOs) accumulate during seed development, and have been thought to be associated with the acquisition of desiccation tolerance (DT) by seeds. Here, comprehensive approaches were adopted to evaluate the changes of DT in developing Arabidopsis seeds of wild type, overexpression (OX-AtGS1/GS2/RS5), and mutant lines by manipulating the expression levels of the GALACTINOL SYNTHASE (GS) and RAFFINOSE SYNTHASE (RS) genes. Our results indicate that seeds of the double mutant (gs1, gs2) and rs5 delayed the timing of DT acquisition as compared to wild type. Subsequent detection confirmed that seeds from OX-AtGS1/GS2 plants with high levels of galactinol, raffinose, and stachyose, and OX-AtRS5 plants possess more raffinose and stachyose but less galactinol compared to wild type. These lines all showed greater germination percentage and shorter time to 50% germination after desiccation treatment at 11 and 15 days after flower (DAF). Further analysis revealed that the role of RFOs is time limited and mainly affects the middle stage (9-16 DAF) of seed development by enhancing seed viability and the ratio of GSH to GSSH in cells, but there is no significant difference in DT of mature seeds. In addition, RFOs could reduce damage to seeds caused by oxidative stress. We conclude that GALACTINOL SYNTHASE and RAFFINOSE SYNTHASE play important roles in DT acquisition during Arabidopsis seed development, and that galactinol and RFOs are crucial protective compounds in the response of seeds to desiccation stress.

Arabidopsis group XIV ubiquitin-conjugating enzymes AtUBC32, AtUBC33, and AtUBC34 play negative roles in drought stress response.

AtUBC32, AtUBC33, and AtUBC34 comprise Arabidopsis group XIV E2 ubiquitin-conjugating enzymes. Yeast two-hybrid, in vitro pull-down, and bimolecular fluorescence complementation assays revealed that group XIV E2s are interacting partners of the U-box-type E3 ligase PUB19, a negative regulator of drought stress response. These three AtUBCs are co-localized with PUB19 to the punctae-like structures, most of which reside on the endoplasmic reticulum membrane of tobacco leaf cells. Suppression of AtUBC32, AtUBC33, and AtUBC34 resulted in increased abscisic acid-mediated stomatal closure and tolerance to drought stress. These results indicate that Arabidopsis group XIV E2s play negative roles in drought stress response.

GmSK1, an SKP1 homologue in soybean, is involved in the tolerance to salt and drought.

In plants, various proteins are regulated by the ubiquitin-mediated system in response to different environmental stresses, such as drought, cold and heat. The Skp1-Cullin-F-box (SCF) complex, one of the multisubunit E3 ligases, has been shown to be involved in abiotic response pathways. In this study, Glycine max SKP1-like 1 (GmSK1), which had the typical characteristics of an SKP1 protein, with an alpha/beta structure, targeted to the cytoplasm and nucleus, was isolated from soybean [Glycine max (L.)]. GmSK1 was constitutively expressed in all the tested tissues, especially in the roots. Furthermore, the expression of GmSK1 was simultaneously induced by abscisic acid (ABA), jasmonic acid (JA), salicylic acid (SA), NaCl, low temperatures and drought, which suggests important roles for GmSK1 in plant responses to hormone treatments and abiotic stress. GmSK1-overexpressing transgenic tobacco (Nicotiana tobacum cv. Samsun) plants showed enhanced tolerance to high salinity and drought stress; exhibited significantly reduced inhibition of growth, greenness and water loss; and exhibited increased MDA accumulation compared with wild-type controls. Our results suggest that GmSK1 might play a role in the crosstalk between ubiquitination and abiotic stress responses in plants.

Exploring the role of Inositol 1,3,4-trisphosphate 5/6 kinase-2 (GmITPK2) as a dehydration and salinity stress regulator in Glycine max (L.) Merr. through heterologous expression in E. coli.

Phytic acid (PA) is implicative in a spectrum of biochemical and physiological processes involved in plant stress response. Inositol 1,3,4, Tris phosphate 5/6 kinase (ITPK), a polyphosphate kinase that converts Inositol 1,3,4 trisphosphate to Inositol 1,3,4,5/6 tetra phosphate, averting the inositol phosphate pool towards PA biosynthesis, is a key regulator that exists in four different isoforms in soybean. In the present study, in-silico analysis of the promoter region of ITPKs was done and among the four isoforms, promoter region of GmITPK2 showed the presence of two MYB binding elements for drought inducibility and one for ABA response. Expression profiling through qRT-PCR under drought and salinity stress showed higher expression of GmITPK2 isoform compared to the other members of the family. The study revealed GmITPK2 as an early dehydration responsive gene which is also induced by dehydration and exogenous treatment with ABA. To evaluate the osmo-protective role of GmITPK2, attempts were made to assess the bacterial growth on Luria Broth media containing 200 mM NaCl, 16% PEG and 100 µM ABA, individually. The transformed E. coli BL21 (DE3) cells harbouring the GmITPK2 gene depicted better growth on the media compared to the bacterial cells containing the vector alone. Similarly, the growth of the transformed cells in the liquid media containing 200 mM NaCl, 16% PEG and 100 µM ABA showed higher absorbance at 600 nm compared to control, at different time intervals. The GmITPK2 recombinant E. coli cells showing tolerance to drought and salinity thus demonstrated the functional redundancy of the gene across taxa. The purity and specificity of the recombinant protein was assessed and confirmed through PAGE showing a band of ∼35 kDa on western blotting using Anti- Penta His- HRP conjugate antibody. To the best of our knowledge, the present study is the first report exemplifying the role of GmITPK2 isoform in drought and salinity tolerance in soybean.

Overexpression of the maize E3 ubiquitin ligase gene ZmAIRP4 enhances drought stress tolerance in Arabidopsis.

Ubiquitin-mediated protein degradation plays a crucial role in enabling plants to effectively and efficiently cope with environmental stresses. The E3 ligases have emerged as a central component of the ubiquitination pathway and modulate plant response to abiotic stresses. However, few such studies have been reported in maize. In this study, a C3HC4-type RING finger E3 ligase in maize, ZmAIRP4 (Zea mays Abscisic acid [ABA]-Insensitive RING Protein 4), which is an ortholog of AtAIRP4, was isolated by reverse transcription polymerase chain reaction with specific primers, and its functions in tolerance to drought stress were described. ZmAIRP4 was upregulated by ABA, polyethylene glycol and sodium chloride. In vitro ubiquitination assays and subcellular localization indicated that ZmAIRP4 was an active E3 ligase predominantly localized in the cytoplasm and nucleus. Compared to wild type, ZmAIRP4-overexpressing Arabidopsis plants were hypersensitive to ABA during early seedling development, and showed enhanced drought tolerance. Moreover, the transcript levels of several drought-related downstream genes in transgenic plants were dramatically increased compared with wild type plants. Our results suggested that E3 ligase ZmAIRP4 is a positive regulator in the drought tolerance response pathway.

A CBL-interacting protein kinase TaCIPK27 confers drought tolerance and exogenous ABA sensitivity in transgenic Arabidopsis.

Drought is one of the major environmental stresses to plants. The calcium sensor, calcineurin B-like (CBL) proteins, and their interacting protein kinases (CIPK) play important roles in responding to abiotic stresses. In this study, we functionally characterized a CIPK gene from Triticum aestivum designated TaCIPK27. The transcriptional levels of TaCIPK27 were increased both in roots and leaves after treatment with polyethylene glycol 8000, abscisic acid and H2O2. Besides, TaCIPK27 interacted with AtCBL1, AtCBL3, AtCBL4, AtCBL5 and AtCBL9 in yeast two-hybrid assays. Ectopic overexpression of TaCIPK27 positively regulates drought tolerance in transgenic Arabidopsis compared with controls, which was demonstrated by seed germination and survival rates experiments, as well as the detection of physiological indices including ion leakage, malonic dialdehyde and H2O2 contents and antioxidant enzyme activities under normal and drought conditions. Moreover, higher concentration of endogenous abscisic acid was detected under drought in TaCIPK27 transgenic plants. In addition, TaCIPK27 transgenic plants were more sensitive to exogenous abscisic acid treatment at seed germination and seedling stage. The expression levels of somedrought stress and abscisic acid related genes were up-regulated in TaCIPK27 transgenic plants. The results suggest that TaCIPK27 functions as a positive regulator under drought partly in an ABA-dependent pathway.

Tardigrades Use Intrinsically Disordered Proteins to Survive Desiccation.

Tardigrades are microscopic animals that survive a remarkable array of stresses, including desiccation. How tardigrades survive desiccation has remained a mystery for more than 250 years. Trehalose, a disaccharide essential for several organisms to survive drying, is detected at low levels or not at all in some tardigrade species, indicating that tardigrades possess potentially novel mechanisms for surviving desiccation. Here we show that tardigrade-specific intrinsically disordered proteins (TDPs) are essential for desiccation tolerance. TDP genes are constitutively expressed at high levels or induced during desiccation in multiple tardigrade species. TDPs are required for tardigrade desiccation tolerance, and these genes are sufficient to increase desiccation tolerance when expressed in heterologous systems. TDPs form non-crystalline amorphous solids (vitrify) upon desiccation, and this vitrified state mirrors their protective capabilities. Our study identifies TDPs as functional mediators of tardigrade desiccation tolerance, expanding our knowledge of the roles and diversity of disordered proteins involved in stress tolerance.

Loss in photosynthesis during senescence is accompanied by an increase in the activity of ß-galactosidase in leaves of Arabidopsis thaliana: modulation of the enzyme activity by water stress.

The precise nature of the developmental modulation of the activity of cell wall hydrolases that breakdown the wall polysaccharides to maintain cellular sugar homeostasis under sugar starvation environment still remains unclear. In this work, the activity of ß-galactosidase (EC 3.2.1.23), a cell-wall-bound enzyme known to degrade the wall polysaccharides, has been demonstrated to remarkably enhance during senescence-induced loss in photosynthesis in Arabidopsis thaliana. The enhancement in the enzyme activity reaches a peak at the terminal phase of senescence when the rate of photosynthesis is at its minimum. Although the precise nature of chemistry of the interface between the decline in photosynthesis and enhancement in the activity of the enzyme could not be fully resolved, the enhancement in its activity in dark and its suppression in light or with exogenous sugars may indicate the involvement of loss of photosynthetic production of sugars as a key factor that initiates and stimulates the activity of the enzyme. The hydrolase possibly participates in the catabolic network of cell wall polysaccharides to produce sugars for execution of energy-dependant senescence program in the background of loss of photosynthesis. Drought stress experienced by the senescing leaves accelerates the decline in photosynthesis with further stimulation in the activity of the enzyme. The stress recovery of photosynthesis and suppression of the enzyme activity on withdrawal of stress support the proposition of photosynthetic modulation of the cell-wall-bound enzyme activity.

Abscinazole-E3M, a practical inhibitor of abscisic acid 8'-hydroxylase for improving drought tolerance.

Abscisic acid (ABA) is an essential phytohormone that regulates plant water use and drought tolerance. However, agricultural applications of ABA have been limited because of its rapid inactivation in plants, which involves hydroxylation of ABA by ABA 8'-hydroxylase (CYP707A). We previously developed a selective inhibitor of CYP707A, (-)-Abz-E2B, by structurally modifying S-uniconazole, which functions as an inhibitor of CYP707A and as a gibberellin biosynthetic enzyme. However, its synthetic yield is too low for practical applications. Therefore, we designed novel CYP707A inhibitors, Abz-T compounds, that have simpler structures in which the 1,2,3-triazolyl ring of (-)-Abz-E2B has been replaced with a triple bond. They were successfully synthesised in shorter steps, resulting in greater yields than that of (-)-Abz-E2B. In the enzymatic assays, one of the Abz-T compounds, (-)-Abz-E3M, acted as a strong and selective inhibitor of CYP707A, similar to (-)-Abz-E2B. Analysis of the biological effects in Arabidopsis revealed that (-)-Abz-E3M enhanced ABA's effects more than (-)-Abz-E2B in seed germination and in the expression of ABA-responsive genes. Treatment with (-)-Abz-E3M induced stomatal closure and improved drought tolerance in Arabidopsis. Furthermore, (-)-Abz-E3M also increased the ABA response in rice and maize. Thus, (-)-Abz-E3M is a more practical and effective inhibitor of CYP707A than (-)-Abz-E2B.

Loss of function of the yellow-e gene causes dehydration-induced mortality of adult Tribolium castaneum.

Yellow protein (dopachrome conversion enzyme, DCE) is involved in the melanin biosynthetic pathway that significantly accelerates pigmentation reactions in insects. Recent studies have suggested that the insect yellow genes represent a rapidly evolving gene family generating functionally diverse paralogs, but the exact physiological functions of several yellow genes are still not understood. To study the function(s) of one of the yellow genes, yellow-e (TcY-e), in the red flour beetle, Tribolium castaneum, we performed real-time PCR to analyze its developmental and tissue-specific expression, and utilized immunohistochemistry to identify the localization of the TcY-e protein in adult cuticle. Injection of double-stranded RNA for TcY-e (dsTcY-e) into late instar larvae had no effect on larval-pupal molting or pupal development. The pupal cuticle, including that lining the setae, gin traps and urogomphi, underwent normal tanning. Adult cuticle tanning including that of the head, mandibles and legs viewed through the translucent pupal cuticle was initiated on schedule (pupal days 4-5), indicating that TcY-e is not required for pupal or pharate adult cuticle pigmentation in T. castaneum. The subsequent pupal-adult molt, however, was adversely affected. Although pupal cuticle apolysis and slippage were evident, some of the adults (~25%) were unable to shed their exuvium and died entrapped in their pupal cuticle. In addition, the resulting adults rapidly became highly desiccated. Interestingly, both the failure of the pupal-adult molt and desiccation-induced mortality were prevented by maintaining the dsTcY-e-treated insects at 100% relative humidity (rh). However, when the high humidity-rescued adults were removed from 100% rh and transferred to 50% rh, they rapidly dehydrated and died, whereas untreated beetles thrived throughout development at 50% rh. We also observed that the body color of the high humidity-rescued dsTcY-e-adults was slightly darker than that of control animals. These results support the hypothesis that TcY-e has a role not only in normal body pigmentation in T. castaneum adults but also has a vital waterproofing function.

Clinical Significance of Fecal Lactoferrin and Multiplex Polymerase Chain Reaction in Patients with Acute Diarrhea.

BACKGROUND/AIMS: The diagnostic yield of fecal leukocyte and stool cultures is unsatisfactory in patients with acute diarrhea. This study was performed to evaluate the clinical significance of the fecal lactoferrin test and fecal multiplex polymerase chain reaction (PCR) in patients with acute diarrhea. METHODS: Clinical parameters and laboratory findings, including fecal leukocytes, fecal lactoferrin, stool cultures and stool multiplex PCR for bacteria and viruses, were evaluated prospectively for patients who were hospitalized due to acute diarrhea. RESULTS: A total of 54 patients were included (male, 23; median age, 42.5 years). Fecal leukocytes and fecal lactoferrin were positive in 33 (61.1%) and 14 (25.4%) patients, respectively. Among the 31 patients who were available for fecal pathogen evaluation, fecal multiplex PCR detected bacterial pathogens in 21 patients, whereas conventional stool cultures were positive in only one patient (67.7% vs 3.2%, p=0.000). Positive fecal lactoferrin was associated with presence of moderate to severe dehydration and detection of bacterial pathogens by multiplex PCR (21.4% vs 2.5%, p=0.049; 100% vs 56.5%, p=0.032, respectively). CONCLUSIONS: Fecal lactoferrin is a useful marker for more severe dehydration and bacterial etiology in patients with acute diarrhea. Fecal multiplex PCR can detect more causative organisms than conventional stool cultures in patients with acute diarrhea.

Clinical Significance of Fecal Lactoferrin and Multiplex Polymerase Chain Reaction in Patients with Acute Diarrhea

BACKGROUND/AIMS: The diagnostic yield of fecal leukocyte and stool cultures is unsatisfactory in patients with acute diarrhea. This study was performed to evaluate the clinical significance of the fecal lactoferrin test and fecal multiplex polymerase chain reaction (PCR) in patients with acute diarrhea. METHODS: Clinical parameters and laboratory findings, including fecal leukocytes, fecal lactoferrin, stool cultures and stool multiplex PCR for bacteria and viruses, were evaluated prospectively for patients who were hospitalized due to acute diarrhea. RESULTS: A total of 54 patients were included (male, 23; median age, 42.5 years). Fecal leukocytes and fecal lactoferrin were positive in 33 (61.1%) and 14 (25.4%) patients, respectively. Among the 31 patients who were available for fecal pathogen evaluation, fecal multiplex PCR detected bacterial pathogens in 21 patients, whereas conventional stool cultures were positive in only one patient (67.7% vs 3.2%, p=0.000). Positive fecal lactoferrin was associated with presence of moderate to severe dehydration and detection of bacterial pathogens by multiplex PCR (21.4% vs 2.5%, p=0.049; 100% vs 56.5%, p=0.032, respectively). CONCLUSIONS: Fecal lactoferrin is a useful marker for more severe dehydration and bacterial etiology in patients with acute diarrhea. Fecal multiplex PCR can detect more causative organisms than conventional stool cultures in patients with acute diarrhea.

Cyclical dehydration-induced renal injury and Mesoamerican nephropathy: as sweet by any other name?

Severe cyclical dehydration induces chronic renal injury in rodents. This effect is attenuated by global fructokinase deficiency, suggesting possible roles for fructokinase and fructose metabolism in mediating or promoting dehydration-induced injury. Clinical and pathological similarities between this injury model and endemic Mesoamerican nephropathy (MeN) have fueled speculation that dehydration-induced injury and MeN may share common mechanistic underpinnings involving fructokinase that can be targeted to mitigate disease development, progression, and/or severity.

A transcriptional approach to unravel the connection between phospholipases A2 and D and ABA signal in citrus under water stress.

The effect of water stress on the interplay between phospholipases (PL) A2 and D and ABA signalling was investigated in fruit and leaves from the sweet orange Navelate and its fruit-specific ABA-deficient mutant Pinalate by studying simultaneously expression of 5 PLD and 3 PLA2-encoding genes. In general, expression levels of PLD-encoding genes were higher at harvest in the flavedo (coloured outer part of the peel) from Pinalate. Moreover, a higher and transient increase in expression of CsPLDα, CsPLDß, CsPLDδ and CsPLDζ was observed in the mutant as compared to Navelate fruit under water stress, which may reflect a mechanism of acclimation to water stress influenced by ABA deficiency. An early induction in CsPLDγ gene expression, when increase in peel damage during fruit storage was most evident, suggested a role for this gene in membrane degradation processes during water stress. Exogenous ABA on mutant fruit modified the expression of all PLD genes and reduced the expression of CsPLDα and CsPLDß by 1 week to levels similar to those of Navelate, suggesting a repressor role of ABA on these genes. In general, CssPLA2α and ß transcript levels were lower in flavedo from Pinalate than from Navelate fruit during the first 3 weeks of storage, suggesting that expression of these genes also depends at least partially on ABA levels. Patterns of expression of PLD and PLA2-encoding genes were very similar in Navelate and Pinalate leaves, which have similar ABA levels, when comparing both RH conditions. Results comparison with other from previous works in the same experimental systems helped to decipher the effect of the stress severity on the differential response of some of these genes under dehydration conditions and pointed out the interplay between PLA2 and PLD families and their connection with ABA signalling in citrus.

Fructokinase activity mediates dehydration-induced renal injury.

The epidemic of chronic kidney disease in Nicaragua (Mesoamerican nephropathy) has been linked with recurrent dehydration. Here we tested whether recurrent dehydration may cause renal injury by activation of the polyol pathway, resulting in the generation of endogenous fructose in the kidney that might subsequently induce renal injury via metabolism by fructokinase. Wild-type and fructokinase-deficient mice were subjected to recurrent heat-induced dehydration. One group of each genotype was provided water throughout the day and the other group was hydrated at night, after the dehydration. Both groups received the same total hydration in 24 h. Wild-type mice that received delayed hydration developed renal injury, with elevated serum creatinine, increased urinary NGAL, proximal tubular injury, and renal inflammation and fibrosis. This was associated with activation of the polyol pathway, with increased renal cortical sorbitol and fructose levels. Fructokinase-knockout mice with delayed hydration were protected from renal injury. Thus, recurrent dehydration can induce renal injury via a fructokinase-dependent mechanism, likely from the generation of endogenous fructose via the polyol pathway. Access to sufficient water during the dehydration period can protect mice from developing renal injury. These studies provide a potential mechanism for Mesoamerican nephropathy.