bluuebunny/crossref_metadata_2025 huggingface dataset:
B-50/GAP-43-induced Formation of Filopodia Depends on Rho-GTPase
Authors: Aarts Lambertus H. J. , Schrama Loes H. , Hage Willem J. , Bos Johannes L. , Gispen Willem Hendrik , Schotman Peter
Published: July 2013

In the present study we show that expression of the neural PKC-substrate B-50 (growth-associated protein [GAP-43]) in Rat-1 fibroblasts induced the formation of filopodial extensions during spreading. This morphological change was accompanied by an enhanced formation of peripheral actin filaments and by accumulation of vinculin immunoreactivity in filopodial focal adhesions, colocalizing with B-50. In time lapse experiments, the B-50–induced filopodial extensions were shown to stay in close contact with the substratum and appeared remarkably stable, resulting in a delayed lamellar spreading of the fibroblasts. The morphogenetic effects of the B-50 protein were entirely dependent on the integrity of the two N-terminal cysteines involved in membrane association (C3C4), but were not significantly affected by mutations of the PKC-phosphorylation site (S41) or deletion of the C terminus (177–226). Cotransfection of B-50 with dominant negative Cdc42 or Rac did not prevent B-50–induced formation of filopodial cells, whereas this process could be completely blocked by cotransfection with dominant negative Rho or Clostridium botulinum C3-transferase. Conversely, constitutively active Rho induced a similar filopodial phenotype as B-50. We therefore propose that the induction of surface extensions by B-50 in spreading Rat-1 fibroblasts depends on Rho-guanosine triphosphatase function.
TheUNI3Gene Is Required for Assembly of Basal Bodies ofChlamydomonasand Encodes δ-Tubulin, a New Member of the Tubulin Superfamily
Authors: Dutcher Susan K. , Trabuco Emanuel C.
Published: July 2013

We have cloned the UNI3 gene inChlamydomonas and find that it encodes a new member of the tubulin superfamily. Although Uni3p shares significant sequence identity with α-, β-, and γ-tubulins, there is a region of Uni3p that has no similarity to tubulins or other known proteins. Mutantuni3–1 cells assemble zero, one, or two flagella. Pedigree analysis suggests that flagellar number inuni3–1 cells is a function of the age of the cell. The uniflagellate uni3–1 cells show a positional phenotype; the basal body opposite the eyespot templates the single flagellum. A percentage of uni3–1 cells also fail to orient the cleavage furrow properly, and basal bodies have been implicated in the placement of cleavage furrows in Chlamydomonas. Finally when uni3–1 cells are observed by electron microscopy, doublet rather than triplet microtubules are observed at the proximal end of the basal bodies. We propose that the Uni3 tubulin is involved in both the function and cell cycle-dependent maturation of basal bodies/centrioles.
RhoG GTPase Controls a Pathway That Independently Activates Rac1 and Cdc42Hs
Authors: Gauthier-Rouvière Cécile , Vignal Emmanuel , Mériane Mayya , Roux Pierre , Montcourier Philippe , Fort Philippe
Published: July 2013

RhoG is a member of the Rho family of GTPases that shares 72% and 62% sequence identity with Rac1 and Cdc42Hs, respectively. We have expressed mutant RhoG proteins fused to the green fluorescent protein and analyzed subsequent changes in cell surface morphology and modifications of cytoskeletal structures. In rat and mouse fibroblasts, green fluorescent protein chimera and endogenous RhoG proteins colocalize according to a tubular cytoplasmic pattern, with perinuclear accumulation and local concentration at the plasma membrane. Constitutively active RhoG proteins produce morphological and cytoskeletal changes similar to those elicited by a simultaneous activation of Rac1 and Cdc42Hs, i.e., the formation of ruffles, lamellipodia, filopodia, and partial loss of stress fibers. In addition, RhoG and Cdc42Hs promote the formation of microvilli at the cell apical membrane. RhoG-dependent events are not mediated through a direct interaction with Rac1 and Cdc42Hs targets such as PAK-1, POR1, or WASP proteins but require endogenous Rac1 and Cdc42Hs activities: coexpression of a dominant negative Rac1 impairs membrane ruffling and lamellipodia but not filopodia or microvilli formation. Conversely, coexpression of a dominant negative Cdc42Hs only blocks microvilli and filopodia, but not membrane ruffling and lamellipodia. Microtubule depolymerization upon nocodazole treatment leads to a loss of RhoG protein from the cell periphery associated with a reversal of the RhoG phenotype, whereas PDGF or bradykinin stimulation of nocodazole-treated cells could still promote Rac1- and Cdc42Hs-dependent cytoskeletal reorganization. Therefore, our data demonstrate that RhoG controls a pathway that requires the microtubule network and activates Rac1 and Cdc42Hs independently of their growth factor signaling pathways.
Effects of Age on the Posttranscriptional Regulation of CCAAT/Enhancer Binding Protein α and CCAAT/Enhancer Binding Protein β Isoform Synthesis in Control and LPS-Treated Livers
Authors: Hsieh Ching-Chyuan , Xiong Wei , Xie Qizhi , Rabek Jeffrey P. , Scott Sheen G. , An Mi Ra , Reisner Peter D. , Kuninger David T. , Papaconstantinou John
Published: July 2013

The CCAAT/enhancer binding protein α (C/EBPα) and CCAAT/enhancer binding protein β (C/EBPβ) mRNAs are templates for the differential translation of several isoforms. Immunoblotting detects C/EBPαs with molecular masses of 42, 38, 30, and 20 kDa and C/EBPβs of 35, 20, and ∼8.5 kDa. The DNA-binding activities and pool levels of p42C/EBPαand p30C/EBPαin control nuclear extracts decrease significantly whereas the binding activity and protein levels of the 20-kDa isoforms increase dramatically with LPS treatment. Our studies suggest that the LPS response involves alternative translational initiation at specific in-frame AUGs, producing specific C/EBPα and C/EBPβ isoform patterns. We propose that alternative translational initiation occurs by a leaky ribosomal scanning mechanism. We find that nuclear extracts from normal aged mouse livers have decreased p42C/EBPαlevels and binding activity, whereas those of p20C/EBPαand p20C/EBPβare increased. However, translation of 42-kDa C/EBPα is not down-regulated on polysomes, suggesting that aging may affect its nuclear translocation. Furthermore, recovery of the C/EBPα- and C/EBPβ-binding activities and pool levels from an LPS challenge is delayed significantly in aged mouse livers. Thus, aged livers have altered steady-state levels of C/EBPα and C/EBPβ isoforms. This result suggests that normal aging liver exhibits characteristics of chronic stress and a severe inability to recover from an inflammatory challenge.
Functional Domains of Rep2, a Transcriptional Activator Subunit for Res2–Cdc10, Controlling the Cell Cycle “Start”
Authors: Tahara Sayaka , Tanaka Koichi , Yuasa Yasuhito , Okayama Hiroto
Published: July 2013

In the fission yeast Schizosaccharomyces pombe, passage from G1to S-phase requires the execution of the transcriptional factor complex that consists of the Cdc10 and Res1/2 molecules. This complex activates the MluI cell cycle box cis-element contained in genes essential for S-phase onset and progression. The rep2+gene, isolated as a multicopy suppressor of a temperature-sensitivecdc10 mutant, has been postulated to encode a putative transcriptional activator subunit for the Res2–Cdc10 complex. To identify the rep2+function and molecularly define its domain organization, we reconstituted the Res2–Cdc10 complex-dependent transcriptional activation in Saccharomyces cerevisiae. Reconstitution experiments, deletion analyses using one and two hybrid systems, and in vivo Res2 coimmunoprecipitation assays show that the Res2–Cdc10 complex itself can recognize but cannot activate MluI cell cycle box without Rep2, and that consistent with its postulated function, Rep2 contains 45-amino acid Res2 binding and 22-amino acid transcriptional activation domains in the middle and C terminus of the molecule, respectively. The functional essentiality of these domains is also demonstrated by their requirement for rescue of the cold-sensitive rep2deletion mutant of fission yeast.
A General RNA-Binding Protein Complex That Includes the Cytoskeleton-associated Protein MAP 1A
Authors: DeFranco Christopher , Chicurel Marina E. , Potter Huntington
Published: July 2013

Association of mRNA with the cytoskeleton represents a fundamental aspect of RNA physiology likely involved in mRNA transport, anchoring, translation, and turnover. We report the initial characterization of a protein complex that binds RNA in a sequence-independent but size-dependent manner in vitro. The complex includes a ∼160-kDa protein that is bound directly to mRNA and that appears to be either identical or highly related to a ∼1600-kDa protein that binds directly to mRNA in vivo. In addition, the microtubule-associated protein, MAP 1A, a cytoskeletal associated protein is a component of this complex. We suggest that the general attachment of mRNA to the cytoskeleton may be mediated, in part, through the formation of this ribonucleoprotein complex.
The Yeast Dynactin Complex Is Involved in Partitioning the Mitotic Spindle between Mother and Daughter Cells during Anaphase B
Authors: Kahana Jason A. , Schlenstedt Gabriel , Evanchuk Darren M. , Geiser John R. , Hoyt M. Andrew , Silver Pamela A.
Published: July 2013

Although vertebrate cytoplasmic dynein can move to the minus ends of microtubules in vitro, its ability to translocate purified vesicles on microtubules depends on the presence of an accessory complex known as dynactin. We have cloned and characterized a novel gene,NIP100, which encodes the yeast homologue of the vertebrate dynactin complex protein p150glued. Like strains lacking the cytoplasmic dynein heavy chain Dyn1p or the centractin homologue Act5p, nip100Δ strains are viable but undergo a significant number of failed mitoses in which the mitotic spindle does not properly partition into the daughter cell. Analysis of spindle dynamics by time-lapse digital microscopy indicates that the precise role of Nip100p during anaphase is to promote the translocation of the partially elongated mitotic spindle through the bud neck. Consistent with the presence of a true dynactin complex in yeast, Nip100p exists in a stable complex with Act5p as well as Jnm1p, another protein required for proper spindle partitioning during anaphase. Moreover, genetic depletion experiments indicate that the binding of Nip100p to Act5p is dependent on the presence of Jnm1p. Finally, we find that a fusion of Nip100p to the green fluorescent protein localizes to the spindle poles throughout the cell cycle. Taken together, these results suggest that the yeast dynactin complex and cytoplasmic dynein together define a physiological pathway that is responsible for spindle translocation late in anaphase.
IκB Is a Substrate for a Selective Pathway of Lysosomal Proteolysis
Authors: Cuervo Ana Maria , Hu Wei , Lim Bing , Dice J. Fred
Published: July 2013

In lysosomes isolated from rat liver and spleen, a percentage of the intracellular inhibitor of the nuclear factor κ B (IκB) can be detected in the lysosomal matrix where it is rapidly degraded. Levels of IκB are significantly higher in a lysosomal subpopulation that is active in the direct uptake of specific cytosolic proteins. IκB is directly transported into isolated lysosomes in a process that requires binding of IκB to the heat shock protein of 73 kDa (hsc73), the cytosolic molecular chaperone involved in this pathway, and to the lysosomal glycoprotein of 96 kDa (lgp96), the receptor protein in the lysosomal membrane. Other substrates for this degradation pathway competitively inhibit IκB uptake by lysosomes. Ubiquitination and phosphorylation of IκB are not required for its targeting to lysosomes. The lysosomal degradation of IκB is activated under conditions of nutrient deprivation. Thus, the half-life of a long-lived pool of IκB is 4.4 d in serum-supplemented Chinese hamster ovary cells but only 0.9 d in serum-deprived Chinese hamster ovary cells. This increase in IκB degradation can be completely blocked by lysosomal inhibitors. In Chinese hamster ovary cells exhibiting an increased activity of the hsc73-mediated lysosomal degradation pathway due to overexpression of lamp2, the human form of lgp96, the degradation of IκB is increased. There are both short- and long-lived pools of IκB, and it is the long-lived pool that is subjected to the selective lysosomal degradation pathway. In the presence of antioxidants, the half-life of the long-lived pool of IκB is significantly increased. Thus, the production of intracellular reactive oxygen species during serum starvation may be one of the mechanisms mediating IκB degradation in lysosomes. This selective pathway of lysosomal degradation of IκB is physiologically important since prolonged serum deprivation results in an increase in the nuclear activity of nuclear factor κ B. In addition, the response of nuclear factor κ B to several stimuli increases when this lysosomal pathway of proteolysis is activated.
ADP-Ribosylation Factors Do Not Activate Yeast Phospholipase Ds but Are Required for Sporulation
Authors: Rudge Simon A. , Cavenagh Margaret M. , Kamath Reena , Sciorra Vicki A. , Morris Andrew J. , Kahn Richard A. , Engebrecht JoAnne
Published: July 2013

ADP-ribosylation factor (ARF) proteins in Saccharomyces cerevisiae are encoded by two genes, ARF1 andARF2. The addition of the c-myc epitope at the C terminus of Arf1 resulted in a mutant (arf1-myc arf2) that supported vegetative growth and rescued cells from supersensitivity to fluoride, but homozygous diploids failed to sporulate. arf1-myc arf2 mutants completed both meiotic divisions but were unable to form spores. The SPO14 gene encodes a phospholipase D (PLD), whose activity is essential for mediating the formation of the prospore membrane, a prerequisite event for spore formation. Spo14 localized normally to the developing prospore membrane in arf1-myc arf2 mutants; however, the synthesis of the membrane was attenuated. This was not a consequence of reduced PLD catalytic activity, because the enzymatic activity of Spo14 was unaffected in meiotic arf1-myc arf2 mutants. Although potent activators of mammalian PLD1, Arf1 proteins did not influence the catalytic activities of either Spo14 or ScPld2, a second yeast PLD. These results demonstrate that ARF1 is required for sporulation, and the mitotic and meiotic functions of Arf proteins are not mediated by the activation of any known yeast PLD activities. The implications of these results are discussed with respect to current models of Arf signaling.
Gap Junctional Communication Modulates Gene Expression in Osteoblastic Cells
Authors: Lecanda Fernando , Towler Dwight A. , Ziambaras Konstantinos , Cheng Su-Li , Koval Michael , Steinberg Thomas H. , Civitelli Roberto
Published: July 2013

Bone-forming cells are organized in a multicellular network interconnected by gap junctions. In these cells, gap junctions are formed by connexin43 (Cx43) and connexin45 (Cx45). Cx43 gap junctions form pores that are more permeable to negatively charged dyes such as Lucifer yellow and calcein than are Cx45 pores. We studied whether altering gap junctional communication by manipulating the relative expression of Cx43 and Cx45 affects the osteoblast phenotype. Transfection of Cx45 in cells that express primarily Cx43 (ROS 17/2.8 and MC3T3-E1) decreased both dye transfer and expression of osteocalcin (OC) and bone sialoprotein (BSP), genes pivotal to bone matrix formation and calcification. Conversely, transfection of Cx43 into cells that express predominantly Cx45 (UMR 106–01) increased both cell coupling and expression of OC and BSP. Transient cotransfection of promoter–luciferase constructs and connexin expression vectors demonstrated that OC and BSP gene transcription was down-regulated by Cx45 cotransfection in ROS 17/2.8 and MC3T3-E1 cells, in association with a decrease in dye coupling. Conversely, cotransfection of Cx43 in UMR 106–01 cells up-regulated OC and BSP gene transcription. Activity of other less specific osteoblast promoters, such as osteopontin and osteonectin, was less sensitive to changes in gap junctional communication. Thus, altering gap junctional permeability by manipulating the expression of Cx43 and Cx45 in osteoblastic cells alters transcriptional activity of osteoblast-specific promoters, presumably via modulation of signals that can diffuse from cell to cell. A communicating intercellular network is required for the full elaboration of a differentiated osteoblastic phenotype.
A Novel Fluorescence-based Genetic Strategy Identifies Mutants ofSaccharomyces cerevisiaeDefective for Nuclear Pore Complex Assembly
Authors: Bucci Mirella , Wente Susan R.
Published: July 2013

Nuclear pore complexes (NPCs) are large proteinaceous portals for exchanging macromolecules between the nucleus and the cytoplasm. Revealing how this transport apparatus is assembled will be critical for understanding the nuclear transport mechanism. To address this issue and to identify factors that regulate NPC formation and dynamics, a novel fluorescence-based strategy was used. This approach is based on the functional tagging of NPC proteins with the green fluorescent protein (GFP), and the hypothesis that NPC assembly mutants will have distinct GFP-NPC signals as compared with wild-type (wt) cells. By fluorescence-activated cell sorting for cells with low GFP signal from a population of mutagenized cells expressing GFP-Nup49p, three complementation groups were identified: two correspond to mutantnup120 and gle2 alleles that result in clusters of NPCs. Interestingly, a third group was a novel temperature-sensitive allele of nup57. The lowered GFP-Nup49p incorporation in the nup57-E17 cells resulted in a decreased fluorescence level, which was due in part to a sharply diminished interaction between the carboxy-terminal truncated nup57pE17and wt Nup49p. Interestingly, thenup57-E17 mutant also affected the incorporation of a specific subset of other nucleoporins into the NPC. Decreased levels of NPC-associated Nsp1p and Nup116p were observed. In contrast, the localizations of Nic96p, Nup82p, Nup159p, Nup145p, and Pom152p were not markedly diminished. Coincidentally, nuclear import capacity was inhibited. Taken together, the identification of such mutants with specific perturbations of NPC structure validates this fluorescence-based strategy as a powerful approach for providing insight into the mechanism of NPC biogenesis.
Selective Perturbation of Apical Membrane Traffic by Expression of Influenza M2, an Acid-activated Ion Channel, in Polarized Madin–Darby Canine Kidney Cells
Authors: Henkel Jennifer R. , Apodaca Gerard , Altschuler Yoram , Hardy Stephen , Weisz Ora A.
Published: July 2013

The function of acidification along the endocytic pathway is not well understood, in part because the perturbants used to modify compartmental pH have global effects and in some cases alter cytoplasmic pH. We have used a new approach to study the effect of pH perturbation on postendocytic traffic in polarized Madin–Darby canine kidney (MDCK) cells. Influenza M2 is a small membrane protein that functions as an acid-activated ion channel and can elevate the pH of the trans-Golgi network and endosomes. We used recombinant adenoviruses to express the M2 protein of influenza virus in polarized MDCK cells stably transfected with the polymeric immunoglobulin (Ig) receptor. Using indirect immunofluorescence and immunoelectron microscopy, M2 was found to be concentrated at the apical plasma membrane and in subapical vesicles; intracellular M2 colocalized partly with internalized IgA in apical recycling endosomes as well as with the trans-Golgi network marker TGN-38. Expression of M2 slowed the rate of IgA transcytosis across polarized MDCK monolayers. The delay in transport occurred after IgA reached the apical recycling endosome, consistent with the localization of intracellular M2. Apical recycling of IgA was also slowed in the presence of M2, whereas basolateral recycling of transferrin and degradation of IgA were unaffected. By contrast, ammonium chloride affected both apical IgA and basolateral transferrin release. Together, our data suggest that M2 expression selectively perturbs acidification in compartments involved in apical delivery without disrupting other postendocytic transport steps.
Differential Expression and Functions of Cortical Myosin IIA and IIB Isotypes during Meiotic Maturation, Fertilization, and Mitosis in Mouse Oocytes and Embryos
Authors: Simerly Calvin , Nowak Grzegorz , de Lanerolle Primal , Schatten Gerald
Published: July 2013

To explore the role of nonmuscle myosin II isoforms during mouse gametogenesis, fertilization, and early development, localization and microinjection studies were performed using monospecific antibodies to myosin IIA and IIB isotypes. Each myosin II antibody recognizes a 205-kDa protein in oocytes, but not mature sperm. Myosin IIA and IIB demonstrate differential expression during meiotic maturation and following fertilization: only the IIA isoform detects metaphase spindles or accumulates in the mitotic cleavage furrow. In the unfertilized oocyte, both myosin isoforms are polarized in the cortex directly overlying the metaphase-arrested second meiotic spindle. Cortical polarization is altered after spindle disassembly with Colcemid: the scattered meiotic chromosomes initiate myosin IIA and microfilament assemble in the vicinity of each chromosome mass. During sperm incorporation, both myosin II isotypes concentrate in the second polar body cleavage furrow and the sperm incorporation cone. In functional experiments, the microinjection of myosin IIA antibody disrupts meiotic maturation to metaphase II arrest, probably through depletion of spindle-associated myosin IIA protein and antibody binding to chromosome surfaces. Conversely, the microinjection of myosin IIB antibody blocks microfilament-directed chromosome scattering in Colcemid-treated mature oocytes, suggesting a role in mediating chromosome–cortical actomyosin interactions. Neither myosin II antibody, alone or coinjected, blocks second polar body formation, in vitro fertilization, or cytokinesis. Finally, microinjection of a nonphosphorylatable 20-kDa regulatory myosin light chain specifically blocks sperm incorporation cone disassembly and impedes cell cycle progression, suggesting that interference with myosin II phosphorylation influences fertilization. Thus, conventional myosins break cortical symmetry in oocytes by participating in eccentric meiotic spindle positioning, sperm incorporation cone dynamics, and cytokinesis. Although murine sperm do not express myosin II, different myosin II isotypes may have distinct roles during early embryonic development.
Coupled Translocation Events Generate Topological Heterogeneity at the Endoplasmic Reticulum Membrane
Authors: Moss Kenneth , Helm Andrew , Lu Yun , Bragin Alvina , Skach William R.
Published: July 2013

Topogenic determinants that direct protein topology at the endoplasmic reticulum membrane usually function with high fidelity to establish a uniform topological orientation for any given polypeptide. Here we show, however, that through the coupling of sequential translocation events, native topogenic determinants are capable of generating two alternate transmembrane structures at the endoplasmic reticulum membrane. Using defined chimeric and epitope-tagged full-length proteins, we found that topogenic activities of two C-trans (type II) signal anchor sequences, encoded within the seventh and eighth transmembrane (TM) segments of human P-glycoprotein were directly coupled by an inefficient stop transfer (ST) sequence (TM7b) contained within the C-terminus half of TM7. Remarkably, these activities enabled TM7 to achieve both a single- and a double-spanning TM topology with nearly equal efficiency. In addition, ST and C-trans signal anchor activities encoded by TM8 were tightly linked to the weak ST activity, and hence topological fate, of TM7b. This interaction enabled TM8 to span the membrane in either a type I or a type II orientation. Pleiotropic structural features contributing to this unusual topogenic behavior included 1) a short, flexible peptide loop connecting TM7a and TM7b, 2) hydrophobic residues within TM7b, and 3) hydrophilic residues between TM7b and TM8.
BNFinder2: Faster Bayesian network learning and Bayesian classification
Authors: Dojer Norbert , Bednarz Paweł , Podsiadło Agnieszka , Wilczyński Bartek
Published: July 2013

Summary: Bayesian Networks (BNs) are versatile probabilistic models applicable to many different biological phenomena. In biological applications the structure of the network is usually unknown and needs to be inferred from experimental data. BNFinder is a fast software implementation of an exact algorithm for finding the optimal structure of the network given a number of experimental observations. Its second version, presented in this article, represents a major improvement over the previous version. The improvements include (i) a parallelized learning algorithm leading to an order of magnitude speed-ups in BN structure learning time; (ii) inclusion of an additional scoring function based on mutual information criteria; (iii) possibility of choosing the resulting network specificity based on statistical criteria and (iv) a new module for classification by BNs, including cross-validation scheme and classifier quality measurements with receiver operator characteristic scores.

Availability and implementation: BNFinder2 is implemented in python and freely available under the GNU general public license at the project Web site https://launchpad.net/bnfinder, together with a user’s manual, introductory tutorial and supplementary methods.

Contact: dojer@mimuw.edu.pl or bartek@mimuw.edu.pl

Supplementary information:  Supplementary data are available at Bioinformatics online.
SbacHTS: Spatial background noise correction for High-Throughput RNAi Screening
Authors: Zhong Rui , Kim Min Soo , White Michael A. , Xie Yang , Xiao Guanghua
Published: June 2013

Motivation: High-throughput cell-based phenotypic screening has become an increasingly important technology for discovering new drug targets and assigning gene functions. Such experiments use hundreds of 96-well or 384-well plates, to cover whole-genome RNAi collections and/or chemical compound files, and often collect measurements that are sensitive to spatial background noise whose patterns can vary across individual plates. Correcting these position effects can substantially improve measurement accuracy and screening success.

Result: We developed SbacHTS (Spatial background noise correction for High-Throughput RNAi Screening) software for visualization, estimation and correction of spatial background noise in high-throughput RNAi screens. SbacHTS is supported on the Galaxy open-source framework with a user-friendly open access web interface. We find that SbacHTS software can effectively detect and correct spatial background noise, increase signal to noise ratio and enhance statistical detection power in high-throughput RNAi screening experiments.

Availability:  http://www.galaxy.qbrc.org/

Contact:  Guanghua.Xiao@UTSouthwestern.edu

Supplementary information:  Supplementary data are available at Bioinformatics online.
“Legitimate commerce” in the Eighteenth Century: The Royal African Company of England Under the Duke of Chandos, 1720–1726
Authors: Mitchell Matthew David
Published: June 2013

Following the loss in 1712 of its previous monopoly over British trade with West Africa, the Royal African Company found itself unable to compete with smaller, lower-cost British slave traders and nearly collapsed entirely. Salvation seemed to arrive in 1720 in the person of James Brydges, the Duke of Chandos, who led a massive re-capitalization of the company and made the strategic decision to move its focus to the commodity trade between Europe and Africa and on the search for new botanical and mineral resources in Africa itself. While Chandos directed the RAC’s employees in implementing this radical new scheme, he kept it secret from his fellow shareholders, leading them to believe that his plans were aimed at revitalizing the company’s mature but declining line of business in the transatlantic slave trade. The Duke’s strategy, however, proved overly ambitious and failed to reverse the company’s decline.
Effect of Milling on the Color, Nutritional Properties, and Antioxidant Contents of Glutinous Black Rice
Authors: Laokuldilok Thunnop , Surawang Suthat , Klinhom Jitra
Published: July 2013

Dehulled glutinous black rice (cv. Kam Doi Saket) was abrasively milled (0–60 s) to make a degree of milling (DOM) curve. The curve showed a nonlinear relationship between milling time and DOM. The slope sharply increased in the first 20 s; after this point, the increase was moderate. The redness of grain and flour surprisingly increased after milling, and the highest value was found at 20 s of milling. Protein, fat, and crude fiber were not uniformly distributed in the dehulled rice kernel. The rice retained only 76.95, 32.79, 20.24, and 36.57% of protein, fat, crude fiber, and ash, respectively, after milling for 60 s. Anthocyanins, γ‐oryzanol, and α‐tocopherol decreased by 74.49, 55.35, and 70.36%, respectively, after 10 s of milling. The scavenging activity decreased sharply when milling was carried out for longer than 30 s. The methanolic extract from rice milled for 60 s showed the lowest activity, which was 21.1 times lower than that of the dehulled rice extract. It was concluded that milling for 20 s was sufficient to remove most of the bran layer of the black rice sample, but 10 s of milling retained higher contents of nutritional components and rice antioxidants.
Removal and Isolation of Germ‐Rich Fractions from Hull‐less Barley Using a Fitzpatrick Comminuting Mill and Sieves
Authors: Moreau Robert A. , Hicks Kevin B.
Published: July 2013

A process was developed to produce a germ‐enriched fraction from hull‐less barley using a Fitzpatrick comminuting mill (FitzMilling) followed by sieving. Hulled and hull‐less barleys contain 1.5–2.5% oil and, like wheat kernels, which contain wheat germ oil, much of the oil in barley kernels is in the germ fraction. A process that combined FitzMilling and sieving produced a germ‐enriched fraction with an oil content of ≈15% and a yield of ≈1.1%. For comparison, this is higher than the levels of oil in most samples of commercial wheat germ. Experimental conditions were also described to produce a germ‐enriched fraction with a higher yield (2.16%), but it would have lower oil content (10.24%). Germination and compositional analysis studies suggested that FitzMilling hull‐less barley for 2 min or longer reduced germination rates to 1% or less, which was interpreted to mean that almost the entire viable germ was removed. In contrast, FitzMilling conventional hulled barley for 4 min had no effect on germination, and milling for 6 and 8 min resulted in germination rates of 36 and 12%, respectively. The oil extracted from germ‐enriched fractions was rich in free phytosterols (≈1%), phytosterol esters (3–7%), and free fatty acids (2–10%). These germ‐enriched fractions and the extracted oil they contain may have value as nutraceuticals or premium edible oils.
Host Cell Entry of Powdery Mildew Is Correlated with Endosomal Transport of Antagonistically Acting VvPEN1 and VvMLO to the Papilla
Authors: Feechan A. , Jermakow A. M. , Ivancevic A. , Godfrey D. , Pak H. , Panstruga R. , Dry I. B.
Published: July 2013

Challenge by a nonadapted powdery mildew fungal pathogen leads to the formation of a local cell-wall apposition (papilla) beneath the point of attempted penetration. Several plasma membrane (PM) proteins with opposing roles in powdery mildew infection, including Arabidopsis thaliana PENETRATION1 (PEN1) and barley (Hordeum vulgare) MILDEW RESISTANCE LOCUS O (MLO), are localized to the site of powdery mildew attack. PEN1 contributes to penetration resistance to nonadapted powdery mildews, whereas MLO is a susceptibility factor required by adapted powdery mildew pathogens for host cell entry. Our previous studies have demonstrated that the vesicle and endosomal trafficking inhibitors, brefeldin A and wortmannin, have opposite effects on the penetration rates of adapted and nonadapted powdery mildews on grapevine. These findings prompted us to study the pathogen-induced intracellular trafficking of grapevine variants of MLO and PEN1. We first identified grapevine (Vitis vinifera) VvPEN1 and VvMLO orthologs that rescue Arabidopsis Atpen1 and Atmlo2 mlo6 mlo12 null mutants, respectively. By using endomembrane trafficking inhibitors in combination with fluorescence microscopy, we demonstrate that VvMLO3/VvMLO4 and VvPEN1 are co-trafficked together from the PM to the site of powdery mildew challenge. This focal accumulation of VvMLO3/VvMLO4 and VvPEN1 to the site of attack seems to be required for their opposing functions during powdery mildew attack, because their subcellular localization is correlated with the outcome of attempted powdery mildew penetration.
First Report of Potato Blackleg Disease Caused by Pectobacterium atrosepticum in Guangdong China
Authors: She X. M. , He Z. F. , Tang Y. F. , Du Z. G. , Lan G. B.
Published: July 2013

Potato (Solanum tuberosum L.) is an important crop in China. In 2013, diseased potatoes exhibiting blackleg and soft rot symptoms were found in the winter potato growing areas of Huizhou city, Guangdong Province, China, with an incidence of approximately 20%. Initially, the stem bases of infected plants blackened and this symptom spread upward. Later, foliage of the diseased plants became yellow and the stem rotted with vascular discoloration. Twenty diseased plants with typical black leg symptoms were collected from a 10-ha potato field with approximately 60,000 potato plants per hectare. A bacterium with small, irregular, round, fluidal, white colonies was isolated from the vascular tissue of all diseased plants on nutrient agar at 26°C for 2 days. Ten strains were randomly selected for pathogenicity assays. Potato plants (cv. Favorita) at the five- to six-leaf stage were inoculated by injecting their stems with 1 ml of each strain in a bacterial suspension (3 × 108 CFU/ml). The inoculated potato plants were incubated at 16 to 21°C and 65 to 85% humidity, and exhibited the same symptoms as the diseased potato plants in the field by 3 to 5 days post inoculation (dpi). The bacterium was reisolated from the diseased tissue (stem) of the inoculated potato plants and produced characteristic pits on crystal violet pectate medium (1). The bacterium utilized a-methyl glucoside, glucose, lactose, maltose, cellobiose, raffinose, melibiose, and citrate, but not d-arabitol, sorbitol, or malonate. The bacteria also gave a positive reaction for catalase and production of reducing substances from sucrose, but gave a negative reaction for oxidase, production of phosphatase, and indole. Using the universal bacterial 16S rDNA primer set, 27f/1541R (4), 1,400-bp fragments were amplified from the 10 strains. The sequences of the 10 fragments (GenBank Accessions KC695819 to KC695828) were identical and had 100% sequence identity with 16S rDNA of Pectobacterium atrosepticum CFBP 1526 (JN600332). Further, the 438-bp and 690-bp fragments were respectively amplified from all 10 strains with the P. atrosepticum-specific primers Y45/Y46 (3) and ECA1f/ECA2r (2). To our knowledge, this is the first report of potato blackleg disease caused by P. atrosepticum (formerly named as Erwinia carotovora subsp. atroseptica) in Guangdong Province, China.

References: (1) D. Cupples et al. Phytopathology 64:468, 1974. (2) S. H. De Boer et al. Phytopathology 85:854, 1995. (3) D. Frenchon et al. Potato Research 41:63, 1995. (4) M. Horita et al. J. Gen. Plant Pathol. 70:278, 2004.
First Report of Charcoal Rot Caused by Macrophomina phaseolina in Soybean in New York
Authors: Cummings J. A. , Bergstrom G. C.
Published: July 2013

Charcoal rot of soybean (Glycine max (L.) Merr.), incited by Macrophomina phaseolina (Tassi) Goidanich, is commonly found in much of the southern soybean production region of the United States, where it can be a major contributor to yield loss in warm, dry seasons (4). The disease has also been reported in northern states including Michigan, Minnesota, North Dakota, and Wisconsin (1,2,3). In early July of the warmer and drier than normal 2012 growing season, wilted soybean plants were observed and collected for diagnosis from a field in Cayuga County, NY. The diseased plants showed brown discoloration of the stem and roots but the diagnostic microsclerotia of M. phaseolina had not yet developed in the epidermis. Cut pieces of roots and crowns were surface-sterilized in 20% sodium hypochlorite, rinsed in sterile distilled water, and incubated on potato dextrose agar at room temperature with a 12-h photoperiod for 7 days. The pathogen was observed growing out of many of the crown and root pieces, and produced abundant microsclerotia as described by Smith and Wyllie (4). Pathogenicity of isolate Mp001NY12 was confirmed using a cut stem inoculation method. Five soybean plants were grown to the unifoliate stage and inoculated by cutting the stem above the unifoliate leaves and applying a fully colonized agar plug to the wound. Within 2 weeks, infection was obvious with microsclerotia formed in the epidermis of all infected plants, and M. phaseolina was reisolated from all infected plants. DNA of isolate Mp001NY12 was extracted from colonies grown on PDA with PrepMan Ultra DNA extraction kit (Applied Biosystems, Foster City, CA), and the DNA was submitted for ITS sequencing with the ITS1 and ITS4 primers used for PCR amplification. The ITS sequence (GenBank Accession No. KC800709) of Mp001NY12 was compared to those in the NCBI GenBank database using a BLAST search, and had 99% nucleotide sequence identity with M. phaseolina (accessions JX945170, FJ415067, and EU250575). To the best of our knowledge, charcoal rot has not been reported previously on soybean or other plant hosts in New York or in other states in the northeastern United States.

References: (1) Baird et al. Mycopathologia 170:169, 2010. (2) C. A. Bradley and L. E. del Rio. Plant Dis. 87:601, 2003. (3) M. E. ElAraby et al. Plant Dis. 87:202, 2003. (4) G. S. Smith and T. D. Wyllie. Charcoal rot. Page 29 in: Compendium of Soybean Diseases, 4th ed. G. L. Hartman et al., eds. APS Press, St. Paul, MN, 1999.
Percolation-Based Risk Index for Pathogen Invasion: Application to Soilborne Disease in Propagation Systems
Authors: Poggi S. , Neri F. M. , Deytieux V. , Bates A. , Otten W. , Gilligan C. A. , Bailey D. J.
Published: July 2013

Propagation systems for seedling growth play a major role in agriculture, and in notable cases (such as organic systems), are under constant threat from soil and seedborne fungal plant pathogens such as Rhizoctonia solani or Pythium spp. Yet, to date little is known that links the risk of disease invasion to the host density, which is an agronomic characteristic that can be readily controlled. We introduce here, for the first time in an agronomic system, a percolation framework to analyze the link. We set up an experiment to study the spread of the ubiquitous fungus R. solani in replicated propagation systems with different planting densities, and fit a percolation-based epidemiological model to the data using Bayesian inference methods. The estimated probability of pathogen transmission between infected and susceptible plants is used to calculate the risk of invasion. By comparing the transmission probability and the risk values obtained for different planting densities, we are able to give evidence of a nonlinear relationship between disease invasion and the inter-plant spacing, hence to demonstrate the existence of a spatial threshold for epidemic invasion. The implications and potential use of our methods for the evaluation of disease control strategies are discussed.
Autophagy is required for exercise training‐induced skeletal muscle adaptation and improvement of physical performance
Authors: Lira Vitor A. , Okutsu Mitsuharu , Zhang Mei , Greene Nicholas P. , Laker Rhianna C. , Breen David S. , Hoehn Kyle L. , Yan Zhen
Published: June 2013

Pathological and physiological stimuli, including acute exercise, activate autophagy; however, it is unknown whether exercise training alters basal levels of autophagy and whether autophagy is required for skeletal muscle adaptation to training. We observed greater autophagy flux (i.e., a combination of increased LC3‐II/LC3‐I ratio and LC3‐II levels and reduced p62 protein content indicating a higher rate of initiation and resolution of autophagic events), autophagy protein expression (i.e., Atg6/Beclin1, Atg7, and Atg8/LC3) and mitophagy protein Bnip3 expression in tonic, oxidative muscle compared to muscles of either mixed fiber types or of predominant glycolytic fibers in mice. Long‐term voluntary running (4 wk) resulted in increased basal autophagy flux and expression of autophagy proteins and Bnip3 in parallel to mitochondrial biogenesis in plantaris muscle with mixed fiber types. Conversely, exercise training promoted autophagy protein expression with no significant increases of autophagy flux and mitochondrial biogenesis in the oxidative soleus muscle. We also observed increased basal autophagy flux and Bnip3 content without increases in autophagy protein expression in the plantaris muscle of sedentary muscle‐specific Pgc‐1α. transgenic mice, a genetic model of augmented mitochondrial biogenesis. These findings reveal that endurance exercise training‐induced increases in basal autophagy, including mitophagy, only take place if an enhanced oxidative phenotype is achieved. However, autophagy protein expression is mainly dictated by contractile activity independently of enhancements in oxidative phenotype. Exercise‐trained mice heterozygous for the critical autophagy protein Atg6 showed attenuated increases of basal autophagy flux, mitochondrial content, and angiogenesis in skeletal muscle, along with impaired improvement of endurance capacity. These results demonstrate that increased basal autophagy is required for endurance exercise training‐induced skeletal muscle adaptation and improvement of physical performance.—Lira, V. A., Okutsu, M., Zhang, M., Greene, N. P., Laker, R. C., Breen, D. S., Hoehn, K. L., Yan, Z., Autophagy is required for exercise training‐induced skeletal muscle adaptation and improvement of physical performance. FASEB J. 27, 4184–4193 (2013). www.fasebj.org
Decreased long‐chain fatty acid oxidation impairs postischemic recovery of the insulin‐resistant rat heart
Authors: Harmancey Romain , Vasquez Hernan G. , Guthrie Patrick H. , Taegtmeyer Heinrich
Published: June 2013

Diabetic patients with acute myocardial infarction are more likely to die than nondiabetic patients. In the present study we examined the effect of insulin resistance on myocardial ischemia tolerance. Hearts of rats, rendered insulin resistant by high‐sucrose feeding, were subjected to ischemia/reperfusion ex vivo. Cardiac power of control hearts from chow‐fed rats recovered to 93%, while insulin‐resistant hearts recovered only to 80% (P< 0.001 vs. control). Unexpectedly, impaired contractile recovery did not result from an impairment of glucose oxidation (576±36 vs. 593±42 nmol/min/g dry weight; not significant), but from a failure to increase and to sustain oxidation of the long‐chain fatty acid oleate on reperfusion (1878±56 vs. 2070±67 nmol/min/g dry weight; P<0.05). This phenomenon was due to a reduced ability to transport oleate into mitochondria and associated with a 38–58% decrease in the mitochondrial uncoupling protein 3 (UCP3) levels. Contractile function was rescued by replacing oleate with a medium‐chain fatty acid or by restoring UCP3 levels with 24 h of food withdrawal. Lastly, the knockdown of UCP3 in rat L6 myocytes also decreased oleate oxidation by 13–18% following ischemia. Together the results expose UCP3 as a critical regulator of long‐chain fatty acid oxidation in the stressed heart postischemia and identify octanoate as an intervention by which myocardial metabolism can be manipulated to improve function of the insulin‐resistant heart.—Harmancey, R., Vasquez, H. G., Guthrie, P. H., Taegtmeyer, H., Decreased long‐chain fatty acid oxidation impairs postischemic recovery of the insulin‐resistant rat heart. FASEB J. 27, 3966–3978 (2013). www.fasebj.org
The use and misuse of photosynthesis in the quest for novel methods to harness solar energy to make fuel
Authors: Cogdell Richard J. , Gardiner Alastair T. , Molina Pedro I. , Cronin Leroy
Published: July 2013

This short review will illustrate that photosynthesis can provide a real contribution towards our sustain- able, green fuel requirements in the future. However, it is argued that the focus on biofuels is misplaced and that, in the longer term, investment in artificial photosynthesis will prove much more beneficial.
Paenalcaligenes hermetiae sp. nov., isolated from the larval gut of Hermetia illucens (Diptera: Stratiomyidae), and emended description of the genus Paenalcaligenes
Authors: Lee Youn Yeop , Lee Jae Kook , Park Kwan Ho , Kim Seo-Yeon , Roh Seong Woon , Lee Sang-Beom , Choi Youngcheol , Lee Sung-Jae
Published: June 2013

A novel Gram-stain-negative, facultatively anaerobic, non-motile and short rod-shaped bacterium, strain KBL009T, was isolated from the larval gut of Hermetia illucens. Strain KBL009T grew optimally at 37 °C, at pH 6.0 and with 1–2 % (w/v) NaCl. The 16S rRNA gene sequence of strain KBL009T showed 97.6 % similarity to that of

Paenalcaligenes hominis

CCUG 53761AT indicating its classification with the genus

Paenalcaligenes

. The major fatty acids were cyclo-C17 : 0, C16 : 0 and summed feature 2 (comprising C14 : 0 3-OH/iso-C16 : 1). The respiratory quinones were ubiquinone-8 (Q-8), predominating, and a minor amount of Q-7. The polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, one unknown aminolipid and five unknown polar lipids. The polyamine pattern contained predominantly putrescine and relatively high amounts of spermidine. The betaproteobacterial-specific 2-hydroxyputrescine could only be detected in trace amounts. The G+C content of genomic DNA was 56.1 mol%. Results from DNA–DNA hybridization with

P. hominis

KCTC 23583T unambiguously demonstrated that strain KBL009T represents a novel species in the genus

Paenalcaligenes

. Based on phenotypic, genotypic and phylogenetic characterization, the novel species Paenalcaligenes hermetiae sp. nov. is proposed. The type strain is KBL009T ( = KACC 16840T = JCM 18423T). An emended description of the genus

Paenalcaligenes

is also provided.