“Pollination” of a fungus by a fly

Summary The mechanism resulting in fertilization of Epichloë typhina, a heterothallic ascomycete that is an endophytic pathogen of grasses, has now been discovered. Conidia of one mating type are produced in stromata and are then transferred by insects to individuals of the opposite mating type. One insect, Phorbia phrenione, is a particularly important vector of conidia. Once conidia of the opposite mating type have been transferred to a stroma, the life cycle continues with the formation of perithecia.     

Liver Preservation by Single Passage Hypothermic “Squirt” Perfusion

A single passage, hypothermic, intermittent squirt perfusion of the pig''s liver is described which has allowed preservation of porcine livers for up to 17 hours. After preservation, orthotopically allografted livers can maintain recipient animals in good health. If this technique can be applied to clinical liver transplantation it should be possible to move livers from one hospital to another in the same way that kidneys are transported. This should increase the availability of donor livers for transplantation and avoid the serious disadvantages of moving the sick recipient just before the operation to the hospital where the donor has died.     

Renaturation of Inactivated Interferons by “Defensive Reversible Denaturation”

Interferon preparations whose activities had been either partially or completely destroyed by a variety of denaturing conditions could be restored to full activity by converting the inactive conformations to linear random coils and “defending” these linear polypeptides with the anionic detergent, sodium dodecyl sulfate, prior to renaturation. Complete restoration of biological activity of both mouse interferons and human fibroblast interferon required that disulfide bonds be reduced prior to renaturation, but partial reactivation was possible without reduction; these data suggest that both mouse and human fibroblast interferon preparations contain distinct molecular species of interferons.     

Eye-specific learning of routes and “signposts” by walking honeybees

This study investigates the honeybee's ability to learn routes based on visual stimuli presented to a single eye, and to then navigate these routes using the other (naive) eye. Bees were trained to walk through a narrow tunnel carrying visual stimuli on the two walls. At the end of the tunnel the bees had to choose between two arms, one of which led to a feeder. In a first experiment, bees had to learn to choose the left arm to get a reward when the right wall carried a yellow grating, but the right arm when the left wall carried a blue grating. The bees learned this task well, indicating that stimuli encountered by different eyes could be associated with different routes. In a second experiment, bees had to turn left when the right eye saw a blue grating, but to the right when the same eye saw a yellow grating. They also learned this task well. In subsequent tests, they chose the correct arm even when these gratings were presented to the untrained eye. These results suggest that there is interocular transfer of route-specific learning with respect to visual stimuli that function as navigational “signposts”. Accepted: 18 December 1997     

Evidence for Nitrogen Fixation by “Dehalococcoides ethenogenes” Strain 195

Genome annotation of the chlorinated ethene-respiring “Dehalococcoides ethenogenes” strain 195 indicated the presence of a complete nitrogenase operon. Here, results from long-term growth experiments, gene expression, and ¹⁵N₂-isotope measurements confirm that strain 195 is capable of fixing atmospheric dinitrogen when a defined fixed-nitrogen source such as ammonium is unavailable.“Dehalococcoides ethenogenes” strain 195 is the first isolated bacterium that is capable of reductively dechlorinating tetrachloroethene and trichloroethene (TCE) to vinyl chloride (VC) and ethene (22). Annotation of the 1.5-Mbp genome of strain 195 has identified 17 intact reductive dehalogenase (RDase) genes (25). The variety of RDases has essentially defined the metabolic capabilities of strain 195 and other Dehalococcoides strains for respiration of chlorinated ethenes (8, 9, 15, 23, 27) and other chlorinated compounds (1, 2, 6, 21), making them important participants in bioremediation processes (19). Expression of different putative RDase genes has been examined previously in pure culture (6) and in Dehalococcoides-containing enrichment cultures (3, 4, 13, 17, 24, 28).Genome annotation of strain 195 has revealed the presence of a nitrogenase-encoding operon (nif) (DET1151-58) typical of those found in anaerobes (25). According to the published genome annotations of four strains of Dehalococcoides, strain 195 is the only one that contains a nif operon (16, 25; Joint Genome Institute, 2009, Integrated Microbial Genomes system [www.jgi.doe.gov]). A nif operon closely related to that in strain 195 has also been identified in a mixed Dehalococcoides-containing community (29); thus, the nitrogen-fixing function might be present in other unsequenced strains of Dehalococcoides.Phylogenetically, the nitrogenase structural genes of strain 195 are clustered with diverse anaerobic Bacteria, including the molybdenum (Mo)-nitrogenase in Clostridium pasteurianum, as well as Archaea, including the Mo-nitrogenase in Methanosarcina barkeri (25, 30). In the genome of strain 195, the presence of an ABC transporter for molybdenum (DET1159-61) and a nifV gene (DET1614), which encodes homocitrate synthetase used in nitrogenase FeMo-cofactor biosynthesis, suggests that the nitrogenase is of the typical molybdenum-iron type (25). While strain 195 is the only sequenced Dehalococcoides isolate that contains a nif operon, Ju et al. (14) previously identified functional nifH genes in dechlorinating organisms from diverse genera such as Sulfurospirillum multivorans, Desulfovibrio dechloracetivorans, and Desulfomonile tiedjei.Aquifers containing groundwater contaminated with chlorinated ethenes can potentially be limited in nutrients. For example, at the Wurtsmith Air Force Base, the chlorinated ethene-contaminated groundwater was found to contain less than 0.09 mM of ammonia, prompting ammonium amendment (26). Little is currently known about the potential effects of nitrogen limitation on reductive dechlorination in the environment, and the demonstration of nitrogen fixation in strain 195 was previously hindered by the use of an undefined medium (21). Here, we present results demonstrating that strain 195 is capable of fixing atmospheric dinitrogen and the physiological implications of the stress caused by nitrogen limitation.     

“Arabidobrassica”: A novel plant obtained by protoplast fusion

Using somatic hybrid cell lines Arabidopsis thaliana+Brassica campestris, obtained by cloning individual protoplast-fusion products as starting material, shoots and flowering plants have been regenerated. Cytological, biochemical, and morphological analyses indicate that genetic material of both species is present in the resultant plants. Shoots and plants obtained from different lines and different regeneration events differed morphologically and genetically. Most regenerants show morphological abnormalities and unusual organizational patterns. Flowering forms have so far been sterile. Asymmetric hybrids (i.e., hybrids bearing most genetic material of one of the parent species and only few chromosomes of the other) were more regular in morphology. The results represent the first case of intergeneric-intertribal hybridization of flowering plants.Dedicated to the memory of G.D. Karpechenko, author of the theory of remote hybridization and creator of Raphanobrassica hybrid (1928)     

Characterization of “Candidatus Liberibacter Asiaticus” Populations by Double-Locus Analyses

“Candidatus Liberibacter asiaticus” (CaLas) is associated with citrus Huanglongbing (HLB, yellow shoot disease), which is highly destructive to world citrus production. Understanding the relationships of CaLas isolates from different geographical regions is important for HLB research and development of disease management strategies. In this study, 301 CaLas isolates [85 Brazil, 132 China, and 84 U.S. (83 Florida and 1 California)] were collected, and genomic variations among them were evaluated based on the analyses of two genomic loci: trn1, characteristic of variable tandem repeat numbers (TRNs), and snp1, characteristic of single nucleotide polymorphisms (SNPs). Locus trn1 revealed the homogeneity of all Brazilian isolates, and locus snp1 revealed the homogeneity of most Florida isolates. When the two loci were analyzed simultaneously, i.e., double-locus (DL) analyses, CaLas isolates were clustered mostly according to geographical origins. DL genotype 1 included 97 % of the Chinese isolates, DL genotype 2 included all Brazilian isolates, and DL genotype 3 included 93 % of the U.S. isolates. DL analyses successfully revealed inter-continental overlapping or movement pattern of CaLas isolates. The isolate recently found in California belonged to Asiatic DL genotype 1.     

The termination of first trimester pregnancy by extraovular “Prostaglandin-impact”

In 30 sedated volunteers first trimester pregnancy had been successfully terminated with Csapo's method of “Prostaglandin-Impact” (1). The patients were 24.1±0.4 years of age, 9.2±0.4 weeks pregnant, para 0.4±0.1. They all received an extraovular dose of 10 mg PG F2α, under intravenous sedation (100 mg pethidium hydrochloride, 50 mg DPP hydrochloride and 0.5 mg atropinum sulfate). Those 15 (out of 30) patients, whose clinical progress was slow received at 8 hours after the first PG Impact (PGI) a 2nd dose of 5 mg PG F2α, increasing the total average dose to 12.5± 0.5 mg.PGI invariably provoked a rapid and high level uterine contracture. At 4 hours after PGI plasma progesterone (P) already decreased by 20% and cyclic intrauterine pressure was in distinct evolution. At 8 hours after PGI, 15 of the 30 patients showed advanced clinical progress and 33% decrease in P levels. These women aborted in 10.2±1.1 hours, when their P levels decreased by 45% (P < 0.001). Abortion was complete in 13 and incomplete in 2 of the patients. The remaining 15 women, whose clinical progress was indistinct at 8 hours after PGI and whose P levels only decreased by 25% received a 2nd dose of 5 mg PG F2α. These women aborted after 18.5±1.5 hours, 9 completely and 6 incompletely, when their P levels decreased (as in the previous group) by 45% (P < 0.001).All the 30 patients aborted after a short IAT of 14.3±1.2 hours. Abortion was complete in 22 women, while 8 retained various amounts of placental tissue. The “Abortion Score” was high, 94.7±1.6. Only 13 patients had side effects, shortly after PGI, manifesting in vomiting and nausea, which were transient and mild. There were no complications during the study and followup. These findings confirm earlier results (1). When supplemented by extensive field trials, the technique of extraovular PGI might be broadly considered for the non-surgical termination of 1st trimester pregnancy.     

Deterministic and stochastic modelling of endosome escape by Staphylococcus aureus: “quorum” sensing by a single bacterium

Deterministic and stochastic models describing quorum sensing by Staphylococcus aureus within an endosome, and the subsequent escape via the production of virulence factors, are developed and analysed. Particular attention is given to a biologically-relevant asymptotic limit of the problem, for which the solutions, including the endosome escape time, can be explicitly characterised in terms of the model parameters.     

Trypanosoma cruzi dihydrolipoamide dehydrogenase is inactivated by myeloperoxidase-generated “reactive species”

Dihydrolipoamide dehydrogenase (LADH) from Trypanosoma cruzi was inactivated by treatment with myeloperoxidase (MPO)-dependent systems. With MPO/H₂O₂/NaCl, LADH lipoamide reductase and diaphorase activities significantly decreased as a function of incubation time. Iodide, bromide, thiocyanide and chloride effectively supplemented the MPO/H₂O₂ system, KI and NaCl being the most and the least effective supplements, respectively. LADH inactivation by MPO/H₂O₂/NaCl and by NaOCl was similarly prevented by thiol compounds such as GSH, L-cysteine, N-acetylcysteine, penicillamine and N-(2-mercaptopropionyl-glycine) in agreement with the role of HOCl in LADH inactivation by MPO/H₂O₂/NaCl. LADH was also inactivated by MPO/NADH/halide, MPO/H₂O₂/NaNO₂ and MPO/NADH/NaNO₂ systems. Catalase prevented the action of the NADH-dependent systems, thus supporting H₂O₂ production by NADH-supplemented LADH. MPO inhibitors (4-aminobenzoic acid hydrazide, and isoniazid), GSH, L-cysteine, L-methionine and L-tryptophan prevented LADH inactivation by MPO/H₂O₂/NaNO₂. Other MPO systems inactivating LADH were (a) MPO/H₂O₂/chlorpromazine; (b) MPO/H₂O₂/monophenolic systems, including L-tyrosine, serotonin and acetaminophen and (c) MPO/H₂O₂/di- and polyphenolic systems, including norepinephrine, catechol, nordihydroguaiaretic acid, caffeic acid, quercetin and catechin. Comparison of the above effects and those previously reported with pig myocardial LADH indicates that both enzymes were similarly affected by the MPO-dependent systems, allowance being made for T. cruzi LADH diaphorase inactivation and the greater sensitivity of its LADH lipoamide reductase activity towards the MPO/H₂O₂/NaCl system and NaOCl.     

“Chromosome exo-skeleton” in plant cells visualized by scanning electron microscopy

Summary Cytoskeletons surrounding the chromosomes of the root tip cells ofPisum sativum and the generative cells ofAllamanda schottii were visualized using Triton X-100 extraction and scanning electron microscopy. The cytoskeleton surrounding the chromosome consisted of a reticulate network of fibres. This is the first report showing the existence of a chromosome exo-skeleton in plant cells.Abbreviations EGTA ethyleneglycol-bis-(-aminoethylether) N,N,N,N-tetraacetic acid - PIPES piperazine-N,N-bis-(2-ethanesulfonic acid)     

“PEST control”: regulation of molecular barcodes by tyrosine phosphatases

The emerging concept of “molecular barcodes” refers to the dynamic combination of post-translational modifications, often of different nature (e.g., phosphorylation and ubiquitination) that gives rise to multiple forms of a protein which can relay distinct signals throughout a cell. In a recent Cell Research paper by Wang et al., the authors report that a PEST domain-containing tyrosine phosphatase, PTPN18, is able to regulate both phosphorylation and ubiquitination of the HER2 oncogene, barcoding HER2 for increased proteasomal degradation rather than for intracellular trafficking.HER2 is a prominent oncogene overexpressed in breast cancer and several other solid cancers, and is a drug target of Trastuzumab, a monoclonal antibody that is FDA-approved for treatment of HER2+ breast cancers¹. HER2 undergoes phosphorylation on several tyrosine residues, each one regulating specific intracellular functions. In a recent paper published in Cell Research, Wang et al.² first defined which phosphorylation sites on HER2 are regulated by protein tyrosine phosphatase non-receptor type 18 (PTPN18). They identified that three HER2 C-terminal tyrosines are dephosphorylated by PTPN18: Y¹¹¹², Y¹¹⁹⁶ and Y¹²⁴⁸. Previous reports have shown that phosphorylation of Y¹¹¹² leads to enhanced Cbl-mediated K63-linked ubiquitination and lysosomal targeting of HER2³, while Y¹¹⁹⁶ phosphorylation leads to increased cell motility and decreased apoptosis^4,5, and Y¹²⁴⁸ phosphorylation activates MAPK signaling⁶. In an elegant series of experiments, and through co-crystallization of PTPN18 with peptides derived from phosphorylated HER2, Wang et al. identified “selective loss-of-function” point mutations of PTPN18 that abolish the ability of the phosphatase to dephosphorylate each one of the three above-mentioned HER2 phosphorylation sites. The authors then proceeded to express these selective loss-of-function PTPN18 mutants in breast cancer cells in order to assess the cellular phenotypes elicited by hyperphosphorylation of each PTPN18-target site. When they expressed the PTPN18 P109A mutant, which is unable to dephosphorylate the pY¹¹¹² site, the authors indeed observed an increase in phosphorylation-dependent HER2 lysosomal trafficking. However, when they subjected cells to complete knockdown of PTPN18, there was a paradoxical increase in HER2 expression. By following this unexpected observation, they identified a new mechanism of PTPN18-mediated regulation of HER2 half-life. They showed that PTPN18 promotes HER2 ubiquitination and proteasomal degradation through a physical interaction between the PTPN18 PEST domain and HER2, which enables the recruitment of the E3 ubiquitin ligase β-Trcp to the HER2 complex. The authors also provided indirect evidence suggesting that phosphorylation of serine residues in the PEST domain of PTPN18 is necessary to promote ubiquitination and degradation of HER2.In addition to unraveling a new phosphorylation- and ubiquitination-based barcoding mechanism of regulation of the intracellular fate of an important oncogene, this study also introduces two novel concepts in the biochemistry and cellular biology of tyrosine phosphatases. First, by providing an example of structure-based design of mutants able to selectively abolish the capacity of dephosphorylating single sites, this study provides rigorous support to the now widely-accepted notion that phosphatases are not promiscuous enzymes. Rather, they recognize different sites in unique ways that depend on structural determinants outside the active site. While this concept is not new, and has led to the successful design of bi-dentate and multi-dentate tyrosine phosphatase inhibitors with enhanced enzyme selectivity features⁷, it is the first time that it is extended to selective targeting of phosphorylation sites within the same molecule. The Wang et al. approach to phosphatase selectivity through structural biology paves the way to the rational design of compounds for selective inhibition of single site dephosphorylation, which would be desirable for biology studies and signaling therapies⁸.The second innovative aspect of the report by Wang et al.² is the demonstration that two domains of a tyrosine phosphatase mediate different post-translational modifications of the same target molecule. Tyrosine phosphatases such as PTPN11 (SHP-2) can regulate signaling transduction through catalytic-dependent and -independent manners⁹. We also recently reported that another PEST-containing tyrosine phosphatase, PTPN22, promotes the ubiquitination of TRAF3 in innate immune receptor signaling through a catalytic activity-independent mechanism¹⁰. The Wang et al. study goes one step forward by showing that PTPN18 can modify phosphorylation and ubiquitination of the same substrate. The phosphorylation-dependent and the ubiquitination-dependent components of HER2 barcoding are respectively regulated by the catalytic domain and the PEST domain of PTPN18. This suggests that one additional feature conferred to tyrosine phosphatases by their characteristic multi-domain organization¹¹ is the ability to modify the same substrate using two different mechanisms.In summary, Wang et al.² present a study of significant scientific breadth on phosphorylation- and ubiquitination-based barcoding of the oncogene HER2 by the tyrosine phosphatase, PTPN18 (Figure 1). A few questions about the molecular biology of this process remain open and warrant further investigation. For example, it remains to be determined what is the binding site on HER2 that interacts with the PEST domain of PTPN18 and the composition of the HER2 complex that enables ubiquitination and proteasomal targeting of HER2. Further investigations are also needed to clarify whether dephosphorylation or other post-translational modifications of the PEST domain of PTPN18 can alter its function, resulting in different barcoding of HER2 and increased HER2 half-life promoted by the catalytic domain of PTPN18. Further investigation of barcoding of HER2 and other signaling molecules by tyrosine phosphatases would be of high relevance for cancer biology and will improve our understanding of tyrosine phosphatases and the ability to selectively target these enzymes for therapy of human diseases.Open in a separate windowFigure 1PTPN18 regulates the tyrosine phosphorylation and ubiquitination barcodes of HER2 at specific residues through its catalytic (PTP) and PEST domains.     

Examining serological diversity of “cowpea” rhizobia by the ELISA technique

To appraise the usefulness of the enzyme-linked immunosorbent assay (ELISA) technique for examining the serological diversity of slow-growing rhizobia, twelve diverse strains from three countries were examined with four antisera. Eleven of the strains were from the cowpea miscellany, and the twelfth was a Rhizobium japonicum strain. Some cowpea strains showed no antigenic relatedness with each other while others were closely related, and some showed a greater affinity with the R. japonicum strain than with other cowpea strains. All of the strains showed antigenic homology to an isolate from a wild Arachis sp., while two strains isolated from adjacent plants of the same cultivar had little homology. These patterns ofrelatedness and diversity clearly demonstrated the utility of the ELISA method, and so it was used to examine 53 strains isolated from cowpeas grown at three West African locations, Maradi (Niger), Ibadan and Onne (Nigeria). Broad ranges of serological diversity were found in the rhizobia at each location, moreover each population had its own general characteristics. Maradi strains were highly reactive with the five antisera used, Onne strains less so, and Ibadan strains even less so. ELISA reactivity correlated with colony morphology but not with nodulation potential.     

“Rational” Management of Dichlorophenols Biodegradation by the Microalga Scenedesmus obliquus

The microalga Scenedesmus obliquus exhibited the ability to biodegrade dichlorophenols (dcps) under specific autotrophic and mixotrophic conditions. According to their biodegradability, the dichlorophenols used can be separated into three distinct groups. Group I (2,4-dcp and 2,6 dcp – no meta-substitution) consisted of quite easily degraded dichlorophenols, since both chloride substituents are in less energetically demanding positions. Group II (2,3-dcp, 2,5-dcp and 3,4-dcp – one meta-chloride) was less susceptible to biodegradation, since one of the two substituents, the meta one, required higher energy for C-Cl-bond cleavage. Group III (3,5-dcp – two meta-chlorides) could not be biodegraded, since both chlorides possessed the most energy demanding positions. In general, when the dcp-toxicity exceeded a certain threshold, the microalga increased the energy offered for biodegradation and decreased the energy invested for biomass production. As a result, the biodegradation per cell volume of group II (higher toxicity) was higher, than group I (lower toxicity) and the biodegradation of dichlorophenols (higher toxicity) was higher than the corresponding monochlorophenols (lower toxicity). The participation of the photosynthetic apparatus and the respiratory mechanism of microalga to biodegrade the group I and the group II, highlighted different bioenergetic strategies for optimal management of the balance between dcp-toxicity, dcp-biodegradability and culture growth. Additionally, we took into consideration the possibility that the intermediates of each dcp-biodegradation pathway could influence differently the whole biodegradation procedures. For this reason, we tested all possible combinations of phenolic intermediates to check cometabolic interactions. The present contribution bring out the possibility of microalgae to operate as “smart” bioenergetic “machines”, that have the ability to continuously “calculate” the energy reserves and “use” the most energetically advantageous dcp-biodegradation strategy. We tried to manipulate the above fact, changing the energy reserves and as a result the chosen strategy, in order to take advantage of their abilities in detoxifying the environment.     

Seasonal variation and an “outbreak” of frog predation by tamarins

We report temporal variation and an “outbreak” of frog predation by moustached tamarins, Saguinus mystax, in north-eastern Peruvian Amazonia. Frog predation rates were generally very low, but strongly increased in October 2015. Other high rates, identified by outlier analyses, were also observed in September–November of other years. Over all study years, predation rates in this 3-month period were significantly higher than those in the remainder of the year, suggesting a seasonal pattern of frog predation by tamarins. Reduced fruit availability or increased frog abundance or a combination of both may be responsible for both the seasonal pattern and the specific “outbreak” of frog predation.     

The game of “gossip” analyzed by the theory of information

This paper deals with an analysis of a distributed model of the game of “gossip”, in which a message is passed through a line of individuals, and the final (in general, garbled) result is compared with the original ungarbled message. The deterioration of information (defined in the sense of Shannon and Wiener) along the line is calculated, and exact as well as asymptotic formulas suggesting approximate linear electric network analogues are obtained.