Nanovesicles Are Secreted during Pollen Germination and Pollen Tube Growth： A Possible Role in Fertilization

Dear Editor, During recent decades, a novel mechanism of secretion has been identified in a wide range of mammalian cells. It involves the release of bioactive membrane nanovesicles （30-100 nm）, termed exosomes, upon the fusion of multivesicular bodies with the plasma membrane （Thery et al., 2009）. Exosomes are implicated in diverse functions, such as scavenging of archaic proteins, intercellular messengers delivering cell-specific signals, and vehicles for transmissible pathogens. Exosomes have also been described in other organisms such as bacte- ria, Drosophila, and fungi.     

TRIM46 activates AKT/HK2 signaling by modifying PHLPP2 ubiquitylation to promote glycolysis and chemoresistance of lung cancer cells

The incidence of lung cancer is increasing worldwide. Although great progress in lung cancer treatment has been made, the clinical outcome is still unsatisfactory. Tripartite motif (TRIM)-containing proteins has been shown to be closely related to tumor progression. However, the function of TRIM46 in lung cancer is largely unknown. Here, TRIM46 amplification was found in lung adenocarcinoma (LUAD) tissues and TRIM46 amplification was significantly associated with a poor survival rate. Overexpression of wild type TRIM46 increased the proliferation of LUAD cells and glycolysis, promoted xenografts growth, and enhanced cisplatin (DDP) resistance of LUAD cells via increased ubiquitination of pleckstrin homology domain leucine-rich repeat protein phosphatase 2 (PHLPP2) and upregulation of p-AKT. In contrast, overexpression of RING-mutant TRIM46 did not show any effects, suggesting the function of TRIM46 was dependent on the E3 ligase activity. Furthermore, we found that TRIM46 promoted LUAD cell proliferation and DDP resistance by enhancing glycolysis. PHLPP2 overexpression reversed the effects of TRIM46 overexpression. Amplification of TRIM46 also promoted LUAD growth and enhanced its DDP resistance in a patient-derived xenograft (PDX) model. In conclusion, our data highlight the importance of TRIM46/PHLPP2/AKT signaling in lung cancer and provide new insights into therapeutic strategies for lung cancer.Subject terms: Cancer, Biomarkers     

Direct detection of beta-1,3-glucanase isozymes on polyacrylamide electrophoresis and isoelectrofocusing gels

A procedure to assay isozymes of beta-1,3-glucanase directly on polyacrylamide gel electrophoresis (PAGE) and isoelectrofocusing (IEF) gels by using 2,3,5-triphenyltetrazolium chloride is described. The reagent reacts with reducing sugars released by beta-1,3-glucanases from the substrate laminarin. Acidic and neutral isozymes of beta-1,3-glucanase were detected and quantified on 17.5% native PAGE gels run with an anodic buffer system. A significant linear relationship (alpha = less than 0.01, R = 0.991) was observed between amounts of beta-1,3-glucanase loaded and intensity of bands stained with the reagent on native PAGE gels. A full isozyme pattern was obtained on 7.5% IEF gels with a pH range of 3.5-9.5. The IEF gels were heated in a microwave oven during the staining process to minimize diffusion.     

Structure and dynamics of small soluble Aβ(1-40) oligomers studied by top-down hydrogen exchange mass spectrometry

Aβ peptides can assemble into amyloid fibrils, which represent one of the hallmarks of Alzheimer's disease. Recent studies, however, have focused on the behavior of small soluble Aβ oligomers that possess a much greater neurotoxicity than mature fibrils. The structural characterization of these oligomers remains difficult because of their highly dynamic and polymorphic nature. This work explores the behavior of Aβ(1-40) in a slightly basic solution (pH 9.3) at a low salt concentration (10 mM ammonium acetate). These conditions lead to the formation of small oligomers, without any signs of fibrillation for several hours. The structure and dynamics of these oligomers were characterized by circular dichroism spectroscopy, size exclusion chromatography, and millisecond time-resolved hydrogen exchange mass spectrometry (MS). Our results reveal rapid interconversion between Aβ(1-40) oligomers and monomers. The mole fraction of monomeric molecules is on the order of 40%. Oligomers consist of ~4 Aβ(1-40) molecules on average, and the resulting assemblies have a predominantly β-sheet secondary structure. Hydrogen exchange proceeds in the EX1 regime. This feature allows the application of conformer-specific top-down MS. Electron capture dissociation is used for interrogating the deuteration behavior of the Aβ(1-40) oligomers. This approach provides a spatial resolution of ~2 residues. The backbone amide deuteration pattern uncovered in this way is consistent with a β-turn-β motif for L17-M35. The N-terminus is involved in hydrogen bonding, as well, whereas protection gradually tapers off for C-terminal residues 35-40. Our data are consistent with earlier proposals, according to which Aβ(1-40) oligomers adopt a β-barrel structure. In general terms, this study demonstrates how top-down MS with precursor ion selection can be employed for structural studies of specific protein conformers within a heterogeneous mix.     

Molecular characterization of avian-like H1N1 swine influenza a viruses isolated in Eastern China, 2011

Currently, three predominant subtypes of influenza virus are prevalent in pig populations worldwide: H1N1, H3N2, and H1N2. European avian-like H1N1 viruses, which were initially detected in European pig populations in 1979, have been circulating in pigs in eastern China since 2007. In this study, six influenza A viruses were isolated from 60 swine lung samples collected from January to April 2011 in eastern China. Based on whole genome sequencing, molecular characteristics of two isolates were determined. Phylogenetic analysis showed the eight genes of the two isolates were closely related to those of the avian-like H1N1 viruses circulating in pig populations, especially similar to those found in China. Four potential glycosylation sites were observed at positions 13, 26, 198, 277 in the HA1 proteins of the two isolates. Due to the presence of a stop codon at codon 12, the isolates contained truncated PB1-F2 proteins. In this study, the isolates contained 591Q, 627E and 701N in the polymerase subunit PB2, which had been shown to be determinants of virulence and host adaptation. The isolates also had a D rather than E at position 92 of the NS1, a marker of mammalian adaptation. Both isolates contained the GPKV motif at the PDZ ligand domain of the 3′ end of the NS1, a characteristic marker of the European avian-like swine viruses since about 1999, which is distinct from those of avian, human and classical swine viruses. The M2 proteins of the isolates have the mutation (S31N), a characteristic marker of the European avian-like swine viruses since about 1987, which may confer resistance to amantadine and rimantadine antivirals. Our findings further emphasize the importance of surveillance on the genetic diversity of influenza A viruses in pigs, and raise more concerns about the occurrence of cross-species transmission events.     

Parasite reliance on its host gut microbiota for nutrition and survival

Studying the microbial symbionts of eukaryotic hosts has revealed a range of interactions that benefit host biology. Most eukaryotes are also infected by parasites that adversely affect host biology for their own benefit. However, it is largely unclear whether the ability of parasites to develop in hosts also depends on host-associated symbionts, e.g., the gut microbiota. Here, we studied the parasitic wasp Leptopilina boulardi (Lb) and its host Drosophila melanogaster. Results showed that Lb successfully develops in conventional hosts (CN) with a gut microbiota but fails to develop in axenic hosts (AX) without a gut microbiota. We determined that developing Lb larvae consume fat body cells that store lipids. We also determined that much larger amounts of lipid accumulate in fat body cells of parasitized CN hosts than parasitized AX hosts. CN hosts parasitized by Lb exhibited large increases in the abundance of the bacterium Acetobacter pomorum in the gut, but did not affect the abundance of Lactobacillus fructivorans which is another common member of the host gut microbiota. However, AX hosts inoculated with A. pomorum and/or L. fructivorans did not rescue development of Lb. In contrast, AX larvae inoculated with A. pomorum plus other identified gut community members including a Bacillus sp. substantially rescued Lb development. Rescue was further associated with increased lipid accumulation in host fat body cells. Insulin-like peptides increased in brain neurosecretory cells of parasitized CN larvae. Lipid accumulation in the fat body of CN hosts was further associated with reduced Bmm lipase activity mediated by insulin/insulin-like growth factor signaling (IIS). Altogether, our results identify a previously unknown role for the gut microbiota in defining host permissiveness for a parasite. Our findings also identify a new paradigm for parasite manipulation of host metabolism that depends on insulin signaling and the gut microbiota.Subject terms: Animal physiology, Microbial ecology