Low rates of iridoid glycoside hydrolysis in two Longitarsus leaf beetles with different feeding specialization confer tolerance to iridoid glycoside containing host plants

Iridoid glycosides are plant defence compounds that are deterrent and/or toxic for unadapted herbivores but are readily sequestered by dietary specialists of different insect orders. Hydrolysis of iridoid glycosides by β‐glucosidase leads to protein denaturation. Insect digestive β‐glucosidases thus have the potential to mediate plant–insect interactions. In the present study, mechanisms associated with iridoid glycoside tolerance are investigated in two closely‐related leaf beetle species (Coleoptera: Chrysomelidae) that feed on iridoid glycoside containing host plants. The polyphagous Longitarsus luridus Scopoli does not sequester iridoid glycosides, whereas the specialist Longitarsus tabidus Fabricius sequesters these compounds from its host plants. To study whether the biochemical properties of their β‐glucosidases correspond to the differences in feeding specialization, the number of β‐glucosidase isoforms and their kinetic properties are compared between the two beetle species. To examine the impact of iridoid glycosides on the β‐glucosidase activity of the generalist, L. luridus beetles are kept on host plants with or without iridoid glycosides. Furthermore, β‐glucosidase activities of both species are examined using an artificial β‐glucosidase substrate and the iridoid glycoside aucubin present in their host plants. Both species have one or two β‐glucosidases with different substrate affinities. Interestingly, host plant use does not influence the specific β‐glucosidase activities of the generalist. Both species hydrolyse aucubin with a much lower affinity than the standard substrate. The neutral pH reduces the β‐glucosidase activity of the specialist beetles by approximately 60% relative to its pH optimum. These low rates of aucubin hydrolysis suggest that the ability to sequester iridoid glycosides has evolved as a key to potentially preventing iridoid glycoside hydrolysis by plant‐derived β‐glucosidases.     

Autolysis of the cell wall. Its possible role in endogenous and IAA-induced growth in epicotyls of Cicer arietinum

Indoleacetic acid (IAA), a factor that induces growth in epicotyls of cicer arietinum L. cv. Castellana, increases the autolytic capacity of the cell walls by 50%, suggesting that autolysis is related to the processes of cell wall loosening that accompany growth. IAA promotes an increase in the specific activities of the enzymes involved in autolysis, mainly α-galactosidase (EC 3.2.1.22). This relationship autolysis-growth. was also observed in a study of the autolytic capacity of cell walls from regions of the epicotyl with different growth capacity. The sugars released and the level of enzymatic protein were higher in the subapical region that towards the base.     

In vitro studies on the subcellular location of glucosidase I and glucosidase II in dog pancreas

When programmed with yeast prepro--factor mRNA, the heterologous reticulocyte/dog pancreas translation system synthesizes two pheromone related polypeptides, a cytosolically located primary translation product (pp--F_cyt, 21 kDa) and a membrane-specific and multiply glycosylated e-factor precursor (pp--F₃, 27.5 kDa). Glycosylation of the membrane specific pp--F₃ species is competitively inhibited by synthetic peptides containing the consensus sequence Asn-Xaa-Thr as indicated by a shift of its molecular mass from 27.5 kDa to about 19.5 kDa (pp--F₀) , whereas the primary translation product pp--Fcyt is not affected. Likewise, only the glycosylated pp--F₃ structure is digested by Endo H yielding a polypeptide with a molecular mass between PP--F₀ and pp--Fcyt. These observations strongly suggest that the primary translation product is proteolytically processed during/on its translocation into the lumen of the microsomal vesicles. We believe that this proteolytic processing is due to the cleavage of a signal sequence from the pp--Fcyt species, although this interpretation contradicts previous data from other groups. The distinct effect exerted by various glycosidase inhibitors (e.g. 1-deoxynojirimycin, N-methyl-dNM, 1-deoxymannojirimycin) on the electrophoretic mobility of the pp--F₃ polypeptide indicates that its oligosaccharide chains are processed to presumbly Man₉-GlcNAc₂ structures under thein vitro conditions of translation. This oligosaccharide processing is most likely to involve the action of glucosidase I and glucosidase II as follows from the specificity of the glycosidase inhibitors applied and the differences of the molecular mass observed in their presence. In addition, several arguments suggest that both trimming enzymes are located in the lumen of the microsomal vesicles derived from endoplasmic reticulum membranes.Abbreviations dNM 1-deoxynojirimycin - N-Me-dNM N-methyl-dNM - dMM 1-deoxymannojirimycin - CCCP carbonylcyanide m-chlorophenyl hydrazone     

Overexpression of the persimmon abscisic acid β‐glucosidase gene (DkBG1) alters fruit ripening in transgenic tomato

β‐Glucosidases (BG) are present in many plant tissues. Among these, abscisic acid (ABA) β‐glucosidases are thought to take part in the adjustment of cellular ABA levels, however the role of ABA‐BG in fruits is still unclear. In this study, through RNA‐seq analysis of persimmon fruit, 10 full‐length DkBG genes were isolated and were all found to be expressed. In particular, DkBG1 was highly expressed in persimmon fruits with a maximum expression 95 days after full bloom (DAFD). We verified that, in vitro, DkBG1 protein can hydrolyze ABA‐glucose ester (ABA‐GE) to release free ABA. Compared with wild‐type, tomato plants that overexpressed DkBG1 significantly upregulated the expression of ABA receptor PYL3/7 genes and showed typical symptoms of ABA hypersensitivity in fruits. DkBG1 overexpression (DkBG1‐OE) accelerated fruit ripening onset by 3–4 days by increasing ABA levels at the pre‐breaker stage and induced early ethylene release compared with wild‐type fruits. DkBG1‐OE altered the expression of ripening regulator NON‐RIPENING (NOR) and its target genes; this in turn altered fruit quality traits such as coloration. Our results demonstrated that DkBG1 plays an important role in fruit ripening and quality by adjusting ABA levels via hydrolysis of ABA‐GE.     

Enzymatic synthesis of perillyl alcohol derivatives and investigation of their antiproliferative activity

The monoterpene perillyl alcohol (POH), an intermediate in the plant terpenoid biosynthetic pathway, has well-established tumor chemopreventive and chemotherapeutic potential. We have previously shown that the primary hydroxyl group of POH is essential for its antitumor and anti-angiogenic activities. In the current study we present the enzymatic synthesis of two POH derivatives with different polar and hydrophobic characteristics, namely perillyl glucoside and perillyl glucoside fatty ester, through a two-step modification. Initial glucosylation of POH on its active hydroxyl group with D-(+)-glucose and subsequent esterification of the perillyl glucoside product with vinyl laurate were carried out using almond β-glucosidase and lipase B from Candida antarctica, respectively, in a low-water system. Optimization of enzymatic reactions was performed to achieve the highest possible conversion yields. The antitumor cell proliferation activity against mouse Lewis lung carcinoma cells was retained in both derivatives, although the perillyl glucoside ester showed greater inhibition than perillyl glucoside. Our results underline the feasibility of enzymatically producing novel bioactive analogs of phytochemicals displaying useful physicochemical properties.     

Meta-analysis supports association of a functional SNP (rs1801133) in the MTHFR gene with Parkinson's disease

The MTHFR is a candidate risk gene for Parkinson's disease (PD), and a functional SNP (rs1801133) in the coding region of this gene has been investigated for the associations with the illness extensively among worldwide populations, but overall the results were inconsistent. Here, to assess the relationship between rs1801133 and risk of PD in general populations, we conducted a systematic meta-analysis by combining all available case–control samples in European and Asian populations, with a total of 1820 PD cases and 7530 healthy controls, and the pooled odds ratios (ORs) and 95% confidence intervals (95% CIs) for rs1801133 and PD were calculated using the Mantel–Haenszel method with a fixed-effect model. Overall, rs1801133 was significantly associated with the risk of PD (allelic model, pooled OR = 1.212 for T allele, 95% CI = 1.097–1.340, p-value = 0.0002). When stratifying for ethnicity, significant association was also observed in European (allelic model, pooled OR = 1.187 for T allele, 95% CI = 1.058–1.332, p-value = 0.004) and Asian samples (allelic model, pooled OR = 1.293 for T allele, 95% CI = 1.058–1.580, p-value = 0.012) respectively. In addition, rs1801133 was also significantly associated with MTHFR mRNA expression in both CEU (European, p-value = 0.0149) and CHB (Chinese, p-value = 0.0178) HapMap populations. Collectively, our meta-analysis suggests that rs1801133 is significantly associated with susceptibility to PD in European and Asian populations, and MTHFR is likely an authentic risk gene for PD.     

High relative air humidity and continuous light reduce stomata functionality by affecting the ABA regulation in rose leaves

Plants developed under high (90%) relative air humidity (RH) have previously been shown to have large, malfunctioning stomata, which results in high water loss during desiccation and reduced dark induced closure. Stomatal movement is to a large extent regulated by abscisic acid (ABA). It has therefore been proposed that low ABA levels contribute to the development of malfunctioning stomata. In this study, we investigated the regulation of ABA content in rose leaves, through hormone analysis and β‐glucosidase quantification. Compared with high RH, rose plants developed in moderate RH (60%) and 20 h photoperiod contained higher levels of ABA and β‐glucosidase activity. Also, the amount of ABA increased during darkness simultaneously as the ABA‐glucose ester (GE) levels decreased. In contrast, plants developed under high RH with 20 h photoperiod showed no increase in ABA levels during darkness, and had low β‐glucosidase activity converting ABA‐GE to ABA. Continuous lighting (24 h) resulted in low levels of β‐glucosidase activity irrespective of RH, indicating that a dark period is essential to activate β‐glucosidase. Our results provide new insight into the regulation of ABA under different humidities and photoperiods, and clearly show that β‐glucosidase is a key enzyme regulating the ABA pool in rose plants.     

Growth, metabolic profiling and enzymes activities of Catharanthus roseus seedlings treated with plant growth regulators

The effect of different growth regulators on growth and the production of terpenoid indole alkaloids as well as some enzymes involved in the biosynthesis were studied in Catharanthus roseus seedlings. The seedlings were grown on MS solid medium containing different concentrations of each growth regulator for a period of one month. Extracted alkaloids were analyzed by HPLC for determination of terpenoid indole alkaloid quantities. Continuous availability of growth regulators induced different alkaloids with variable effects among the regulators. Gibberellic acid at concentration of either 5.8 M or 11.6 M resulted in elongation of shoots with lowering the number of leaves. Abscisic acid has a retardant effect on growth. Ethylene did not effect the growth pattern at concentration of 100 M but seedlings were not tolerant to higher concentrations. Methyljasmonate reduced the growth of the root system. Methyljasmonate was a general inducer for all alkaloids and increased the activity of strictosidine glucosidase. Ethylene applications promoted the pathways towards ajmalicine, serpentine, tabersonine and vindoline. Similar effect as for ethylene was observed for abscisic acid. Salicylic acid treatment increased the production of serpentine, tabersonine and higher concentration of salicylic acid induced vindoline accumulation. Peroxidase activity was also induced by salicylic acid. Gibberellic acid has little effect on alkaloid levels.