The glycine max xylem sap and apoplast proteome

Molecular signaling interactions in the plant apoplast are important for defense and developmental responses. We examined the soybean proteome of the apoplastic conduit of root-to-shoot communication, the xylem stream, using gel electrophoresis combined with two types of tandem mass spectrometry. We examined soybeans for the presence of a Bradyrhizobium japonicum-induced, long distance developmental signal that controls autoregulation of nodulation (AON) to determine if xylem proteins (XPs) were involved directly or indirectly in AON. The xylem and apoplast fluids collected in hypocotyl, epicotyl, and stem tissue contained a highly similar set of secreted proteins. The XPs were different from those secreted from imbibing seed implying they play important basic roles in xylem function. The XPs of wild-type and nts1007 plants were indistinguishable irrespective of plant age, inoculation status, or time after inoculation suggesting that none was directly involved in AON. XPs were continuously loaded into the xylem stream, as they were present even 28 h after shoot decapitation. These results were consistent with semiquantitative RT-PCR studies that examined the expression of genes corresponding to the XPs under inoculated or uninoculated conditions. Monitoring the expression of XP genes by RT-PCR showed that four possessed root biased expression. This suggested that the corresponding protein products could be produced in roots and travel long distances to shoots. Of these, a species of lipid transfer protein is a candidate for a water-soluble, long-distance signal-carrier due to the presence of hydrophobic clefts that bind known plant signals in vitro. Two soybean XPs identified in this study, lipid transfer protein and Kunitz trypsin inhibitor (KTI), have known roles in plant signaling.     

Proteomic analysis of microvesicles derived from human colorectal cancer cells

Microvesicles (MV) are membrane vesicles secreted from the plasma and endosomal membrane compartment by various cell types such as hematopoietic, epithelial, and tumor cells. Actively growing tumor cells shed MV, and the rate of shedding increases in malignant tumors. Although recent progress in this area has revealed that tumor-derived MV play multiple roles in tumor growth and metastasis via immune escape, tumor invasion, and angiogenesis, the mechanism of vesicle formation and the biological roles of tumor-derived MV are not understood. Here, we report the first global proteomic analysis of highly purified MV from human colorectal cancer cells. Using 1D SDS gel electrophoresis and nano-LC-MS/MS analyses, we identified a total of 547 microvesicular proteins from three independent experiments with high confidence; 416 proteins were identified at least in two trials, including 181 as yet unreported proteins. We identified 49 proteins involved in the biogenesis of MV, including annexins, ADP-ribosylation factors, and Rab proteins. We also identified 28 proteins that may function in tumorigenesis via promotion of migration, invasion, and growth of tumor cells, immune modulation, metastasis, and angiogenesis. Taken together with previously reported results, our observations suggest that tumor-derived MV may act as communicasomes, that is, extracellular organelles that play diverse roles in intercellular communication. This information will help elucidate the biogenesis and functions of tumor-derived MV, and aid in the development of effective vaccines for various cancers, including colorectal cancer.     

miR-4286 is Involved in Connections Between IGF-1 and TGF-β Signaling for the Mesenchymal Transition and Invasion by Glioblastomas

Cellular and Molecular Neurobiology - The insulin-like growth factor (IGF)-1 and transforming growth factor (TGF)-β signal pathways are both recognized as important in regulating cancer...     

Spatiotemporal distribution and the passive dispersal of fungal spores through HVAC systems

Heating, ventilation and air conditioning (HVAC) systems are widely used to regulate indoor temperature and air quality of modern buildings. The central supply and exhaust system cause the dispersal and removal of bioaerosols. This study presents results from ex situ experiments conducted to a better understanding of the spatiotemporal distribution, passive dispersal and removal of fungal spores through HVAC systems. The study was conducted in a 50 square meters by 3 m high instrument room in a research building with HVAC systems. Plates with Aspergillus flavus colony were attached upside down on the edge of the ceiling vent to imitate the mildew growing. Fungal spores were released for 10 min, collected and counted by the settle plate method in 2 h. Results show that A. flavus spores dispersed to 3.6 m in 2 min and were evenly distributed in the room within 8 min. The concentration of spores decreased from 48 to 3–6 CFU/plate after 60 min and was lower than 1% after 120 min. Our results suggest that airborne fungal spores disperse and remove by HVAC systems efficiently. Without the external and internal source, the HVAC system could greatly reduce the fungal amount in the indoor air to the background level within one hour. This study provided the observed data of the transmission and retention of internal or external biological contaminants through HVAC system.

     

Scarless Genome Editing and Stable Inducible Expression Vectors for Geobacter sulfurreducens

Metal reduction by members of the Geobacteraceae is encoded by multiple gene clusters, and the study of extracellular electron transfer often requires biofilm development on surfaces. Genetic tools that utilize polar antibiotic cassette insertions limit mutant construction and complementation. In addition, unstable plasmids create metabolic burdens that slow growth, and the presence of antibiotics such as kanamycin can interfere with the rate and extent of Geobacter biofilm growth. We report here genetic system improvements for the model anaerobic metal-reducing bacterium Geobacter sulfurreducens. A motile strain of G. sulfurreducens was constructed by precise removal of a transposon interrupting the fgrM flagellar regulator gene using SacB/sucrose counterselection, and Fe(III) citrate reduction was eliminated by deletion of the gene encoding the inner membrane cytochrome imcH. We also show that RK2-based plasmids were maintained in G. sulfurreducens for over 15 generations in the absence of antibiotic selection in contrast to unstable pBBR1 plasmids. Therefore, we engineered a series of new RK2 vectors containing native constitutive Geobacter promoters, and modified one of these promoters for VanR-dependent induction by the small aromatic carboxylic acid vanillate. Inducible plasmids fully complemented ΔimcH mutants for Fe(III) reduction, Mn(IV) oxide reduction, and growth on poised electrodes. A real-time, high-throughput Fe(III) citrate reduction assay is described that can screen numerous G. sulfurreducens strain constructs simultaneously and shows the sensitivity of imcH expression by the vanillate system. These tools will enable more sophisticated genetic studies in G. sulfurreducens without polar insertion effects or need for multiple antibiotics.     

Characterization of a flavin-containing monooxygenase from <Emphasis Type="Italic">Corynebacterium glutamicum</Emphasis> and its application to production of indigo and indirubin

Objective

To examine the role of a gene encoding flavin-containing monooxygenase (cFMO) from Corynebacterium glutamicum ATCC13032 when cloned and expressed in Escherichia coli for the production of indigo pigments.

Results

The blue pigments produced by recombinant E. coli were identified as indigo and indirubin. The cFMO was purified as a fused form with maltose-binding protein (MBP). The enzyme was optimal at 25 °C and pH 8. From absorption spectrum analysis, the cFMO was classified as a flavoprotein. FMO activity was strongly inhibited by 1 mM Cu²⁺ and recovered by adding 1–10 mM EDTA. The enzyme catalyzed the oxidation of TMA, thiourea, and cysteamine, but not glutathione or cysteine. MBP-cFMO had an indole oxygenase activity through oxygenation of indole to indoxyl. The recombinant E. coli produced 685 mg indigo l^?1 and 103 mg indirubin l^?1 from 2.5 g l-tryptophan l^?1.

Conclusion

The results suggest the cFMO can be used for the microbial production of both indigo and indirubin.

     

Large seasonal variation in phytoplankton production in the Amundsen Sea

Polar Biology - To better estimate annual primary production in the Amundsen Sea, which is one of the highest productivity regions in the Southern Ocean, the seasonal variations in carbon and...