Insect attraction to synthetic herbivore‐induced plant volatile‐treated field crops

• 1 Plants produce natural enemy‐attracting semiochemicals known as herbivore‐induced plant volatiles (HIPV) in response to herbivore damage. Deployment of synthetic HIPV in crops could enhance the biological control of pests. To test this, six HIPV [methyl salicylate (MeSA), methyl anthranilate (MeA), methyl jasmonate (MeJA), benzaldehyde (Be), cis‐3‐hexenyl acetate (HA), cis‐hexen‐1‐ol (He)] in three concentrations (0.5%, 1.0% and 2.0% v/v) mixed with a vegetable oil adjuvant, Synertrol® (Organic Crop Protectants Pty Ltd, Australia), were sprayed onto winegrape, broccoli and sweet corn plants.
• 2 The relative abundance of insects within treated plots was assessed with non‐attracting, transparent sticky traps at varying time intervals up to 22 days after spraying.
• 3 In the vineyard experiment, Trichogrammatidae responded to Be and MeA (0.5%) and Be (1.0%); Encyrtidae and Bethylidae responded to MeA (1.0%); Scelionidae responded to all compounds at 1.0% and 2.0%; and predatory insects responded to MeA. In sweet corn, parasitoids as a group and Encyrtidae responded to MeA (0.5%); Braconidae responded to all compounds at 0.5% and Synertrol‐only; thrips responded to all compounds at 0.5% and 1.0%; while all parasitoids responded to all compounds at 0.5% and 1.0% and Synertrol‐only. In broccoli, parasitoids as a group and Scelionidae responded to Be, HA, He and Synertrol‐only; Trichogrammatidae responded to Be (0.5%), He (0.5% and 1.0%), MeJA (1.0%) and MeSA (0.5%); and thrips responded to all compounds at to 0.5% and 1.0%.
• 4 Significant attraction of insects occurred up to 6 days after the HIPV application, suggesting that plants may have been induced to produce endogenous volatiles that attracted insects over an extended period.
• 5 The results obtained are discussed in relation to the potential utility of synthetic HIPV to enhance the biological control of pests.

     

Characterization of an Escherichia coli rotA mutant, affected in periplasmic peptidyl-prolyl cis/trans isomerase

The rotA gene of Escherichia coli encodes a peptidyl-prolyl cis/trans isomerase (PPlase), which is supposed to catalyse protein folding in the periplasm. To investigate the importance of the enzyme, the rotA gene was cloned and a chromosomal deletion mutant was created. The rotA mutant was normally viable. No residual PPlase activity could be detected in the periplasmic fraction of the mutant. Comparison of the patterns of periplasmic and outer membrane proteins by SDS-PAGE revealed no differences in protein composition between the rotA mutant and its parental strain. Similarly, the kinetics of periplasmic protein folding and outer membrane protein assembly appeared unaffected by the rotA mutation. Our results show that the periplasmic PPlase of E. coli is not essential and that the protein does not play an important role in protein folding.     

Determinants of the high-methionine trait in wild and exotic germplasm may have escaped selection during early cultivation of maize

The 18 kDa high-methionine δ-class zein gene from maize has been cloned, and its regulation, structure, and map position studied. These studies have shown that (i) zein genes may also contain tryptophan and lysine codons, (ii) the 18 kDa and the related 10 kDa zein gene are coordinately regulated, but their products accumulate to different levels in a genotype-dependent manner, (iii) the duplication of δ-zein genes probably involved unequal crossing over, (iv) no copy correction in either direction has occurred from teosinte to modern corn, and (v) the duplication of of the 18 kDa zein gene probably occurred before the tetraploidization of a progenitor chromosome. The work shows that important nutritional quality determinants like the high-methionine seed proteins are abundant in several exotic and wild corn varieties and low in most of the inbreds screened. The lack of a selectable phenotype for such quality traits during initial domestication and breeding of corn would have eliminated cis and trans regulatory determinants from the germplasm used in modern corn breeding. Examples of the high-methionine δ-class zeins shown here may be generally applicable in explaining the low nutritional quality of most present-day corn grown.     

Depletion of Kinesin 5B Affects Lysosomal Distribution and Stability and Induces Peri-Nuclear Accumulation of Autophagosomes in Cancer Cells

Background

Enhanced lysosomal trafficking is associated with metastatic cancer. In an attempt to discover cancer relevant lysosomal motor proteins, we compared the lysosomal proteomes from parental MCF-7 breast cancer cells with those from highly invasive MCF-7 cells that express an active form of the ErbB2 (ΔN-ErbB2).

Methodology/Principal Findings

Mass spectrometry analysis identified kinesin heavy chain protein KIF5B as the only microtubule motor associated with the lysosomes in MCF-7 cells, and ectopic ΔN-ErbB2 enhanced its lysosomal association. KIF5B associated with lysosomes also in HeLa cervix carcinoma cells as analyzed by subcellular fractionation. The depletion of KIF5B triggered peripheral aggregations of lysosomes followed by lysosomal destabilization, and cell death in HeLa cells. Lysosomal exocytosis in response to plasma membrane damage as well as fluid phase endocytosis functioned, however, normally in these cells. Both HeLa and MCF-7 cells appeared to express similar levels of the KIF5B isoform but the death phenotype was weaker in KIF5B-depleted MCF-7 cells. Surprisingly, KIF5B depletion inhibited the rapamycin-induced accumulation of autophagosomes in MCF-7 cells. In KIF5B-depleted cells the autophagosomes formed and accumulated in the close proximity to the Golgi apparatus, whereas in the control cells they appeared uniformly distributed in the cytoplasm.

Conclusions/Significance

Our data identify KIF5B as a cancer relevant lysosomal motor protein with additional functions in autophagosome formation.     

Priming effect as determined by adding 14C-glucose to modified controlled composting test

The development of new biodegradable packaging materials, especially biodegradable plastics, has created a need for biodegradability testing. The European standard for controlled composting test was used in this study for assessing if the addition of a test material results in excess CO₂ production in compost. This effect, designated as the priming effect, would give an erroneous result for biodegradation, which is based on CO₂ formation from the test material. Glucose was selected as a test substrate because it is the degradation product of starch and cellulose, which are major compounds of many packaging materials. Both ¹⁴C-glucose and non-labelled glucose was applied to nine compost samples of variable stability and agefrom two weeks to 1.5 years. CO₂ and ¹⁴CO₂ evolution were measured during the incubation. Biodegradation of glucose in unstable composts (age leq6 months) was negative and ¹⁴CO₂ evolution was poor, although the respective composts without glucose produced relatively high amounts of CO₂. It was concluded that a negative priming effect was observed in unstable composts, in which glucose remained mostly non-degraded and apparently inhibited the mineralization of native organic matter in the compost. In stable composts (age 6 months), biodegradation of glucose was high and approximately equal to ¹⁴C-glucose mineralization, i.e., the composts showed no priming effect. Young composts were unsuitable for controlled composting test due to lack of stability. It is important to ensure that the compost inoculum used for the test is sufficiently stable.     

Spatiotemporal Developmental Trajectories in the Arabidopsis Root Revealed Using High-Throughput Single-Cell RNA Sequencing

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Peptidylarginine Deiminase 3 (PAD3) Is Upregulated by Prolactin Stimulation of CID-9 Cells and Expressed in the Lactating Mouse Mammary Gland

Peptidylarginine deiminases (PADs) post-translationally convert arginine into neutral citrulline residues. Our past work shows that PADs are expressed in the canine and murine mammary glands; however, the mechanisms regulating PAD expression and the function of citrullination in the normal mammary gland are unclear. Therefore, the first objective herein was to investigate regulation of PAD expression in mammary epithelial cells. We first examined PAD levels in CID-9 cells, which were derived from the mammary gland of mid-pregnant mice. PAD3 expression is significantly higher than all other PAD isoforms and mediates protein citrullination in CID-9 cells. We next hypothesized that prolactin regulates PAD3 expression. To test this, CID-9 cells were stimulated with 5 μg/mL of prolactin for 48 hours which significantly increases PAD3 mRNA and protein expression. Use of a JAK2 inhibitor and a dominant negative (DN)-STAT5 adenovirus indicate that prolactin stimulation of PAD3 expression is mediated by the JAK2/STAT5 signaling pathway in CID-9 cells. In addition, the human PAD3 gene promoter is prolactin responsive in CID-9 cells. Our second objective was to investigate the expression and activity of PAD3 in the lactating mouse mammary gland. PAD3 expression in the mammary gland is highest on lactation day 9 and coincident with citrullinated proteins such as histones. Use of the PAD3 specific inhibitor, Cl4-amidine, indicates that PAD3, in part, can citrullinate proteins in L9 mammary glands. Collectively, our results show that upregulation of PAD3 is mediated by prolactin induction of the JAK2/STAT5 signaling pathway, and that PAD3 appears to citrullinate proteins during lactation.     

Enhanced error-prone RCA mutagenesis by concatemer resolution

Error-prone rolling circle amplification (RCA) is a promising alternative to error-prone PCR for random mutagenesis. The main disadvantage of error-prone RCA is the low transformation efficiency of the DNA concatemer produced in the amplification reaction. We improved the method by introducing loxP recombination site of bacteriophage P1 Cre recombinase into the target plasmid and reducing the concatemer by Cre recombinase to plasmid-sized units, increasing the number of transformants 50-fold in non-error-prone and 13-fold in error-prone conditions. The efficiency improvement was verified by obtaining 115 ± 57 ceftazidime resistant colonies per recombined RCA reaction from randomly mutated TEM-1 β-lactamase gene library whereas only 9 ± 11 colonies were gained without recombination. Supplementation of the error-prone RCA with Cre/loxP recombination is a simple and useful tool to increase the transformable library size.