The biosynthesis of female moth sex pheromone blends is controlled by a number of different enzymes, many of which are encoded by members of multigene families. One such multigene family, the acyl-CoA desaturases, is composed of certain genes that function as key players in moth sex pheromone biosynthesis. Although much is known regarding the function of some of these genes, very little is known regarding how novel genes have evolved within this family and how this might impact the establishment of new sex pheromone blends within a species. We have discovered that several cryptic ⌬11 and ⌬14 desaturase genes exist in the genomes of the European and Asian corn borers (Ostrinia nubilalis and Ostrinia furnacalis, respectively). Furthermore, an entirely novel class of desaturase gene has arisen in the Ostrinia lineage and is derived from duplication of the ⌬11 desaturase gene and subsequent fusion with a retroposon. Interestingly, the genes have been maintained over relatively long evolutionary time periods in corn borer genomes, and they have not been recognizably pseudogenized, suggesting that they maintain functional integrity. The existence of cryptic desaturase genes in moth genomes indicates that the evolution of moth sex pheromone desaturases in general is much more complex than previously recognized.
Parasitism genes encode effector proteins that are secreted through the stylet of root-knot nematodes to dramatically modify selected plant cells into giant-cells for feeding. The Mi8D05 parasitism gene previously identified was confirmed to encode a novel protein of 382 amino acids that had only one database homolog identified on contig 2374 within the Meloidogyne hapla genome. Mi8D05 expression peaked in M. incognita parasitic second-stage juveniles within host roots and its encoded protein was limited to the subventral esophageal gland cells that produce proteins secreted from the stylet. Constitutive expression of Mi8D05 in transformed Arabidopsis thaliana plants induced accelerated shoot growth and early flowering but had no visible effects on root growth. Independent lines of transgenic Arabidopsis that expressed a double-stranded RNA complementary to Mi8D05 in host-derived RNA interference (RNAi) tests had up to 90% reduction in infection by M. incognita compared with wild-type control plants, suggesting that Mi8D05 plays a critical role in parasitism by the root-knot nematode. Yeast two-hybrid experiments confirmed the specific interaction of the Mi8D05 protein with plant aquaporin tonoplast intrinsic protein 2 (TIP2) and provided evidence that the Mi8D05 effector may help regulate solute and water transport within giant-cells to promote the parasitic interaction.
Oncomirs are microRNAs (miRNA) associated with carcinogenesis and malignant transformation. They have emerged as potential molecular targets for anti-cancer therapy. We hypothesize that grape seed procyanidin extract (GSE) exerts antineoplastic effects through modulations of oncomirs and their downstream targets. We found that GSE significantly down-regulated oncomirs miR-19a and -19b in a variety of lung neoplastic cells. GSE also increased mRNA and protein levels of insulin-like growth factor II receptor (IGF-2R) and phosphatase and tensin homolog (PTEN), both predicted targets of miR-19a and -19b. Furthermore, GSE significantly increased PTEN activity and decreased AKT phosphorylation in A549 cells. Transfection of miR-19a and -19b mimics reversed the up-regulations of IGF2R and PTEN gene expression and abrogated the GSE induced anti-proliferative response. Additionally, oral administration of leucoselect phytosome, comprised of standardized grape seed oligomeric procyanidins complexed with soy phospholipids, to athymic nude mice via gavage, significantly down-regulated miR-19a, -19b and the miR-17-92 cluster host gene (MIR17HG) expressions, increased IGF-2R, PTEN, decreased phosphorylated-AKT in A549 xenograft tumors, and markedly inhibited tumor growth. To confirm the absorption of orally administered GSE, plasma procyanidin B1 levels, between 60 and 90 min after gavage of leucoselect phytosome (400 mg/kg), were measured by LC/MS at week 2 and 8 of treatment; the estimated concentration that was associated with 50% growth inhibition (IC50) (1.3 μg/mL) in vitro was much higher than the IC50 (0.032-0.13 μg/ml) observed in vivo. Our findings reveal novel antineoplastic mechanisms by GSE and support the clinical translation of leucoselect phytosome as an anti-neoplastic and chemopreventive agent for lung cancer.
Meloidogyne incognita is the most economically important plant‐parasitic nematode. Meloidogyne incognita manipulates plant cell development and metabolism by injecting effectors from the oesophageal glands into the plant host. Chorismate mutase (CM) is one such effector that may contribute to successful parasitism by M. incognita. This investigation identified and functionally characterized a novel CM effector, Mi‐CM‐3, which is more similar to CMs from bacteria than from other phytoparasitic nematodes. Spatial and temporal expression assays showed Mi‐cm‐3 mRNA accumulates specifically in the subventral oesophageal glands and transcription is up‐regulated during the early parasitic stages of the nematode. In planta gene silencing of Mi‐cm‐3 attenuated nematode parasitism. In addition, Mi‐cm‐3 could fully restore the full virulence phenotype of the pathogenic bacterium Xanthomonas oryzae pv. oryzae by complementation when it was introduced into a mutant strain carrying a deletion in the CM gene. Transient expression of Mi‐CM‐3 caused a reduction in levels of salicylic acid (SA) and mRNA of gene PR1 in Nicotiana benthamiana in response to oomycete pathogen Phytophthora capsici infection, while confocal observations showed that Mi‐CM‐3 was localized within the cytoplasm and the nucleus, but not the plastids, of transfected N. benthamiana leaf cells. Constitutive expression of Mi‐CM‐3 in N. benthamiana plants inhibited root growth and increased susceptibility to M. incognita infection. These results provide direct experimental evidence to show that Mi‐CM‐3 may play an important role in suppressing plant immunity by regulating the SA pathway during the early parasitic stage of M. incognita so as to promote nematode parasitism.
Grape seed procyanidin extract (GSE) has been reported to exert antineoplastic properties via the inhibition of cyclooxygenase-2 (COX-2)/prostaglandin E 2 (PGE 2 ) eicosanoid pathways. In addition, ample data link carcinogenesis to inflammatory events involving other major eicosanoid metabolic pathways, including prostacyclin (PGI 2 ) and 15-hydroxyeicosatetraenoic acid (15-HETE). We therefore evaluated the effects of GSE on prostacyclin synthase (PTGIS)/PGI 2 and 15-lipoxigenase-2 (15-LOX-2)/15-HETE productions by human lung premalignant and malignant cells and correlated the findings with antiproliferative or proapoptotic effects of GSE. The effects of GSE on PGI 2 and 15-HETE productions by human bronchoalveolar lavage (BAL) cells ex vivo were also determined. We further evaluated the bioactivity of oral administration of leucoselect phytosome (a standardized GSE) in the lungs of subjects participating in a lung cancer chemoprevention trial, by comparing the antiproliferative effects of coculturing matched pre-versus posttreatment BAL fluids with lung premalignant and malignant cells. GSE significantly increased PGI 2 (as measured by 6-keto PGF1a) and 15-HETE productions by these cells. Transfections of PTGIS or 15-LOX-2-specific siRNA partially abrogated the antiproliferative or proapoptotic effects of GSE in lung premalignant and malignant cells, respectively. GSE also increased PTGIS and inhibition of caspase-3, and transfection of 15-LOX-2 siRNA abrogated the GSEinduced apoptosis in A549 cells. In addition, culture supernatants from ex vivo GSE-treated baseline BAL cells, as well as BAL fluids from subjects treated with leucoselect phytosome, significantly decreased proliferations of lung premalignant and malignant cells. Our findings support the continued investigation of GSE as an anti-neoplastic and chemopreventive agent against lung cancer. Cancer Prev Res; 9(12); 925-32. Ó2016 AACR.
The polymerase chain reaction (PCR) was employed to develop a specific assay for plant pathogenic mycoplasmalike organisms (MLOs). A cloned fragment of a plasmid from a severe strain of western aster yellows (AY)-MLO was sequenced to identify oligonucleotide primers for PCR. Amplified DNA fragments of the predicted size were obtained from DNA extracted from plants and insects infected with pear decline MLO, beet leafhopper-transmitted virescence agent, elm yellows MLO and several AY-MLO strains. No amplification occurred from healthy leafhopper or plant DNA. The PCR-based assay was over 500 times more sensitive than a hybridisation-based assay which utilised a cloned AY plasmid fragment as a probe. With the PCR-based assay, MLOs could be detected using DNA samples of leafhoppers that were only crushed and boiled in buffer. Amplification of the target DNA was confirmed by digestion of the PCR product with Mbo I which yielded predicted sized fragments for all MLO strains except Bradford AY and eastern AY. Sequencing the PCR product from elm yellows and eastern AY-MLOs revealed greater than 90% homology, and the failure to restrict the PCR product with Mbo I was due to a single base change in the restriction endonuclease site. The ability of the assay to detect a wide range of MLOs with minimal sample preparation and high sensitivity will be useful in epidemiological studies of MLO-caused diseases.
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