The Frequency of Germline BRCA and Non-BRCA HR-Gene-Variants in a Cohort of Pancreatic Cancer Patients.

Germline DNA alterations affecting homologous recombination pathway genes have been associated with pancreatic cancer (PC) risk. BRCA2 is the most studied gene and affects the management of PC patients and their families. Even though recent reports have suggested a similar role of germline ATM pathogenic variants (PV) in familial PC, there is still a disagreement between experts on how it could affect patient management given the lack of proper PC risk estimates. We retrospectively analyzed the germline data of 257 PC patients among whom nearly 50% were sporadic cases. We showed similar frequencies of BRCA2 (4.9%) and ATM (4.4%) PV or likely pathogenic variants, which were not related to familial history. Based on our findings and that of the literature, we suggest including ATM gene among the panel of genes analyzed in PC patients pending the publication of prospective studies.

Potential use of biochar, compost and iron grit associated with Trifolium repens to stabilize Pb and As on a multi-contaminated technosol.

Vegetation cover can be used in the phytomanagement of polluted areas by adding value to abandoned sites and reducing the dispersion of pollutants by erosion. Appropriate amendments, that allow both efficient plant growth and the immobilization of contaminants in the soil must be chosen in order to optimize the efficiency of this process. We used a mining technosol mainly contaminated by arsenic (1068 mg kg-1) and lead (23387 mg kg-1) to study the effect of three amendments (biochar, compost and iron grit) on (i) physico-chemical properties of the soil and soil pore water, (ii) metal(loid) mobility, bioavailability and bioaccessibility (CaCl2 and Simple Bioaccessibility Extraction Test (SBET)), and (iii) the capability of Trifolium repens to germinate and grow. All the amendments used increased the pH and electrical conductivity of the SPW, resulting in a 90% decrease in the concentration of lead in the soil pore water (SPW). We also demonstrated a decrease in Pb phytoavailability. The amendments allowed the establishment of a plant cover, although the addition of iron grit alone did not allow any clover germination. For the Pontgibaud technosol, the combination of the three amendments resulted in a significant decrease in As and Pb concentrations in clover tissues, mainly in the aerial organs. The amendments also made it possible for some of them to halve the phytoavailable fraction of arsenic. However, for compost, both the As concentrations in the SPW, and the bioavailable fraction of As increased. All the amendments used had contrasting effects on the bioaccessible fractions of metal(loid)s. The most efficient amendment combination was the addition of 5% biochar and 5% compost.

Capability of amendments (biochar, compost and garden soil) added to a mining technosol contaminated by Pb and As to allow poplar seed (Populus nigra L.) germination.

The germination capacity of poplar seeds has never been studied in the context of metal(loid)-contaminated soils, even though poplars are present over a vast geographical area. In this study, black poplar seeds from the Loire Valley (France) were grown for 28 days in mesocosm on a heavily polluted soil that was subjected to different amendments. This phytomanagement process aimed to allow the revegetation of an As and Pb-contaminated mining soil by adding appropriate amendments, resulting in metal(loid) soil stabilisation and efficient plant growth. The objectives were to evaluate the effect of three amendments (garden soil, compost and biochar) when added alone or combined to a technosol on (i) the soil physicochemical properties, (ii) the mobility of As and Pb in the soil pore water (SPW), (iii) the capacity of poplar seeds to germinate and to grow and (iv) the metal(loid) distribution within the plant organs. The addition of amendments alone or combined allowed a 90% decrease in SPW Pb concentrations, while the arsenic concentrations were between 18 and 416 times higher. However, we were only able to obtain seed germination and plant growth on amended soils. These promising results will allow us to explore the use of such amendments in rehabilitating areas that are sources of significant metal(loid) dissemination, as well as allowing a natural plant recolonisation of these sites by seeds from the surrounding environment.

Litsea cubeba leaf essential oil from Vietnam: chemical diversity and its impacts on antibacterial activity.

The threat of bacterial resistance to antibiotics has created an urgent need to develop new antimicrobials. The aim of this study was to characterize the chemical diversity of Litsea cubeba leaf essential oil (EO) and its impacts on the antibacterial activity against pathogenic bacteria. Essential oils collected from seven provinces in North Vietnam (n = 25) were characterized by their high content in either 1,8-cineole or linalool. Linalool-type EOs were more effective against the eight bacterial strains tested than 1,8-cineole-type. Oil samples, LC19 (50% 1,8-cineole) and BV27 (94% linalool), were selected to investigate their antibacterial mechanisms against Escherichia coli. A strong bactericidal effect was observed after 4 and 2 h of exposure respectively. Microscopic analysis of treated E. coli cultures clearly showed that EOs caused changes in cell morphology, loss of integrity and permeability of the cell membrane, as well as DNA loss. However, the effects of both EOs were distinct. LC19 mostly affected cell membrane, led to a significant cell filamentation rate and altered cell width, whereas BV27 damaged cell membrane integrity leading to cell permeabilization and altered nucleoid morphology with the appearance of spot and visibly altered compaction. SIGNIFICANCE AND IMPACT OF THE STUDY: This study aimed to characterize the chemical diversity of Litsea cubeba leaf essential oil (EO) and its impacts on its antibacterial activity. Two major chemotypes (1,8-cineole or linalool rich) were identified in North Vietnam and both were bactericidal against several pathogenic bacteria. A distinct inhibitory effect of EO samples on Escherichia coli was observed. 1,8-cineole-rich sample (LC19) affected cell membrane, led to cell filamentation and perturbation of cell width, while the linalool-rich one (BV27) induced damages in the cell membrane and changes in the nucleoid morphology. The study demonstrates the importance of considering chemotype variations in terms of chemical composition as well as the mode of action.

Antibacterial activity of Litsea cubeba (Lauraceae, May Chang) and its effects on the biological response of common carp Cyprinus carpio challenged with Aeromonas hydrophila.

AIMS: The aims of this study were to characterize the antibacterial activity and the chemotype of Litsea cubeba leaf essential oil (EO) harvested in North Vietnam and to investigate the biological effects induced by the leaf powder on growth, nonspecific immunity and survival of common carp (Cyprinus carpio) challenged with Aeromonas hydrophila. METHODS AND RESULTS: The EO showed the prevalence of linalool (95%, n = 5). It was bactericidal against the majority of tested strains, with minimum inhibitory concentrations ranging from 0·72 to 2·89 mg ml(-1) (Aer. hydrophila, Edwarsiella tarda, Vibrio furnissii, Vibrio parahaemolyticus, Streptococcus garvieae, Escherichia coli, Salmonella Typhimurium). The fish was fed with 0 (control), 2, 4 and 8% leaf powder supplementation diets for 21 days. Nonspecific immunity parameters (lysozyme, haemolytic and bactericidal activities of plasma) were assessed 21 days after feeding period and before the experimental infection. Weight gain, specific growth rate and feed conversion ratio were improved by supplementation of L. cubeba in a dose-related manner, and a significant difference appeared at the highest dose (8%) when compared to the control. The increase in plasma lysozyme was significant for all the treated groups. Haemolysis activity was higher for the groups fed with 4 and 8% plant powder. Antibacterial activity increased significantly for the 8% dose only. CONCLUSIONS: Litsea cubeba leaf powder increased nonspecific immunity of carps in dose-related manner. After infection with Aer. hydrophila, survivals of fish fed with 4 and 8% L. cubeba doses were significantly higher than those fed with 2% dose and the control. SIGNIFICANCE AND IMPACT OF THE STUDY: A range of 4-8% L. cubeba leaf powder supplementation diet (from specific linalool-rich chemotype) can be used in aquaculture to reduce antibiotic burden and impacts of diseases caused by Aer. hydrophila.

N-terminal palmitoylation is required for Toxoplasma gondii HSP20 inner membrane complex localization.

Toxoplasma gondii is an obligate intracellular parasite and the causative agent of toxoplasmosis. Protein palmitoylation is known to play roles in signal transduction and in enhancing the hydrophobicity of proteins thus contributing to their membrane association. Global inhibition of protein palmitoylation has been shown to affect T. gondii physiology and invasion of the host cell. However, the proteins affected by this modification have been understudied. This paper shows that the small heat shock protein 20 from T. gondii (TgHSP20) is synthesized as a mature protein in the cytosol and is palmitoylated in three cysteine residues. However, its localization at the inner membrane complex (IMC) is dependent only on N-terminal palmitoylation. Absence or incomplete N-terminal palmitoylation causes TgHSP20 to partially accumulate in a membranous structure. Interestingly, TgHSP20 palmitoylation is not responsible for its interaction with the daughter cells IMCs. Together, our data describe the importance of palmitoylation in protein targeting to the IMC in T. gondii.

Resolving the polyphyletic nature of Pyricularia (Pyriculariaceae).

Species of Pyricularia (magnaporthe-like sexual morphs) are responsible for major diseases on grasses. Pyricularia oryzae (sexual morph Magnaporthe oryzae) is responsible for the major disease of rice called rice blast disease, and foliar diseases of wheat and millet, while Pyricularia grisea (sexual morph Magnaporthe grisea) is responsible for foliar diseases of Digitaria. Magnaporthe salvinii, M. poae and M. rhizophila produce asexual spores that differ from those of Pyricularia sensu stricto that has pyriform, 2-septate conidia produced on conidiophores with sympodial proliferation. Magnaporthe salvinii was recently allocated to Nakataea, while M. poae and M. rhizophila were placed in Magnaporthiopsis. To clarify the taxonomic relationships among species that are magnaporthe- or pyricularia-like in morphology, we analysed phylogenetic relationships among isolates representing a wide range of host plants by using partial DNA sequences of multiple genes such as LSU, ITS, RPB1, actin and calmodulin. Species of Pyricularia s. str. belong to a monophyletic clade that includes all P. oryzae/P. grisea isolates tested, defining the Pyriculariaceae, which is sister to the Ophioceraceae, representing two novel families. These clades are clearly distinct from species belonging to the Gaeumannomyces pro parte/Magnaporthiopsis/Nakataea generic complex that are monophyletic and define the Magnaporthaceae. A few magnaporthe- and pyricularia-like species are unrelated to Magnaporthaceae and Pyriculariaceae. Pyricularia oryzae/P. grisea isolates cluster into two related clades. Host plants such as Eleusine, Oryza, Setaria or Triticum were exclusively infected by isolates from P. oryzae, while some host plant such as Cenchrus, Echinochloa, Lolium, Pennisetum or Zingiber were infected by different Pyricularia species. This demonstrates that host range cannot be used as taxonomic criterion without extensive pathotyping. Our results also show that the typical pyriform, 2-septate conidium morphology of P. grisea/P. oryzae is restricted to Pyricularia and Neopyricularia, while most other genera have obclavate to more ellipsoid 2-septate conidia. Some related genera (Deightoniella, Macgarvieomyces) have evolved 1-septate conidia. Therefore, conidium morphology cannot be used as taxonomic criterion at generic level without phylogenetic data. We also identified 10 novel genera, and seven novel species. A re-evaluation of generic and species concepts within Pyriculariaceae is presented, and novelties are proposed based on morphological and phylogenetic data.

Effectiveness of SplashGuard Caregiver prototype in reducing the risk of aerosol transmission in intensive care unit rooms of SARS-CoV-2 patients: a prospective and simulation study.

BACKGROUND: The contagiousness of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is known to be linked to the emission of bioaerosols. Thus, aerosol-generating procedures (AGPs) could increase the risk of infection among healthcare workers (HCWs). AIM: To investigate the impact of an aerosol protection box, the SplashGuard Caregiver (SGGC) with suction system, by direct analysis of the presence of viral particles after an AGP, and by using the computational fluid dynamics (CFD) simulation method. METHODS: This prospective observational study investigated HCWs caring for patients with SARS-CoV-2 admitted to an intensive care unit (ICU). Rooms were categorized as: SGCG present and SGCG absent. Virus detection was performed through direct analysis, and using a CFD model to simulate the movement dynamics of airborne particles produced by a patient's respiratory activities. FINDINGS: Of the 67 analyses performed, three samples tested positive on quantitative polymerase chain reaction: one of 33 analyses in the SCCG group (3%) and two of 34 analyses in the non-SGCG group (5.9%). CFD simulations showed that: (1) reduction of the gaps of an SGCG could decrease the number of emitted particles remaining airborne within the room by up to 70%; and (2) positioning HCWs facing the opposite direction to the main air flow would reduce their exposure. CONCLUSIONS: This study documented the presence of SARS-CoV-2 among HCWs in a negative pressure ICU room of an infected patient with or without the use of an SGCG. The simulation will help to improve the design of the SGCG and the positioning of HCWs in the room.

Bayesian latent class analysis of the characteristics of diagnostic tests to assess the passive immunity transfer status in neonatal Belgian Blue beef calves.

The aim of the current study was to assess the diagnostic characteristics of radial immunodiffusion (RID), capillary electrophoresis (CE) and digital brix refractometry (Bx) for the diagnosis of failure of passive transfer (FPT) of immunity in neonatal Belgian Blue beef calves in the absence of a gold standard using a Bayesian latent class model. Belgian blue beef calves (n = 202) from a large farm in the south of Belgium were blood-sampled at 48-72 h of age and tested for FPT. The true prevalence of FPT in this population of calves was 34.5 % (95 % BCI: 26.1-44.3) using a FPT cut-off point of 10 g IgG/L. This true prevalence increased to 66.3 (95 % BCI: 56.9-74.8) and 88.9 % (95 % BCI: 83.1-93.2) at FPT cut-off points of respectively 18 and 25 g IgG/L serum. The Bland-Altman plot comparing the RID and CE methods, revealed that the serum IgG concentrations obtained by RID were on average 2.25 (95 % CI 1.62-2.88) g/L higher than those measured by CE. Optimal cut-off values for CE, corresponding to the FPT values as measured by RID of 10, 18, and 25 g IgG/L serum, were respectively 10, 15, and 20 g IgG/L. The overall diagnostic accuracy of the three diagnostic tests was comparable at the FPT cut-off point of 10 g IgG/L serum (i.e. 85 %). At higher cut-off points for FPT, the RID and CE assays presumably performed better that the Bx method. In conclusion, we demonstrated that: (1) the CE method is a good alternative for the RID assay, the latter having important constraints when considering its practicality, and (2) the Bx method is a cheap and user-friendly indirect method to evaluate the FPT in new-born Belgian Blue beef calves.

Transfer kinetics of labeled aroma compounds from liquid media into coffee beans during simulated wet processing conditions.

The transfer kinetics of three labelled compounds (butanal, 2-phenyethanol, isoamyl acetate) was studied from a liquid medium into the coffee beans during simulated wet processing using four media (M) (M1: contained dehulled beans, M2: contained demucilaginated beans, M3: contained depulped beans, M4: contained depulped beans with yeast). Trials were carried out at 25 °C, under agitation and for five time periods (0, 6, 12, 24 and 48 h), and then the labelled volatiles were analyzed by SPME-GC-MS. The three labelled molecules were transferred into the coffee beans with different mass transfer rates; reaching at 12hrs in the M4, 0.2 ± 0.03, 11.2 ± 0.66 and 1.3 ± 0.04 µg/g of coffee respectively for butanal, 2-phenyethanol and isoamyl acetate. The parchment resistance significantly affected the mass transfer of the 2-phenylethanol. Butanal and isoamyl acetate underwent metabolic reactions, which decreased their amount in the coffee beans. Furthermore, an interaction between molecules and the yeast was observed and decreased significantly the butanal's transfer.

Assessing the performance of single-copy genes for recovering robust phylogenies.

Phylogenies involving nonmodel species are based on a few genes, mostly chosen following historical or practical criteria. Because gene trees are sometimes incongruent with species trees, the resulting phylogenies may not accurately reflect the evolutionary relationships among species. The increase in availability of genome sequences now provides large numbers of genes that could be used for building phylogenies. However, for practical reasons only a few genes can be sequenced for a wide range of species. Here we asked whether we can identify a few genes, among the single-copy genes common to most fungal genomes, that are sufficient for recovering accurate and well-supported phylogenies. Fungi represent a model group for phylogenomics because many complete fungal genomes are available. An automated procedure was developed to extract single-copy orthologous genes from complete fungal genomes using a Markov Clustering Algorithm (Tribe-MCL). Using 21 complete, publicly available fungal genomes with reliable protein predictions, 246 single-copy orthologous gene clusters were identified. We inferred the maximum likelihood trees using the individual orthologous sequences and constructed a reference tree from concatenated protein alignments. The topologies of the individual gene trees were compared to that of the reference tree using three different methods. The performance of individual genes in recovering the reference tree was highly variable. Gene size and the number of variable sites were highly correlated and significantly affected the performance of the genes, but the average substitution rate did not. Two genes recovered exactly the same topology as the reference tree, and when concatenated provided high bootstrap values. The genes typically used for fungal phylogenies did not perform well, which suggests that current fungal phylogenies based on these genes may not accurately reflect the evolutionary relationships among species. Analyses on subsets of species showed that the phylogenetic performance did not seem to depend strongly on the sample. We expect that the best-performing genes identified here will be very useful for phylogenetic studies of fungi, at least at a large taxonomic scale. Furthermore, we compare the method developed here for finding genes for building robust phylogenies with previous ones and we advocate that our method could be applied to other groups of organisms when more complete genomes are available.

The expression of Clostridium perfringens consensus beta2 toxin is associated with bovine enterotoxaemia syndrome.

Clostridium perfringens has been implicated in a broad array of enteric infections including the fatal haemorrhagic enteritis/enterotoxaemia syndrome in cattle. The beta2 toxin (CPB2), encoded by cpb2, is suspected to be implicated in this syndrome. However, among C. perfringens isolates from cattle suspected of clostridial disease, an atypical allele was recently found to predominate at the cpb2 locus and atypical corresponding CPB2 proteins were shown to be poorly expressed, thus arguing against a biologically significant role of the beta2 toxin in clostridial diseases in cattle. This study compared genotype and phenotype of the beta2 toxin between C. perfringens isolates from a group of healthy calves (n=14, 87 isolates) and from a group of enterotoxaemic calves (n=8, 41 isolates). PCR results revealed the exclusive presence of the typical "consensus"cpb2 in the enterotoxaemic group. Western blot analysis demonstrated that the typical variant of CPB2 was often expressed in isolates from enterotoxaemic calves (43.9%) and infrequently in isolates from healthy cattle (6.9%). These data suggest that the typical variant of the CPB2 toxin may play a role in the pathogenesis of cattle enterotoxaemia.

Sublethal effects of epoxiconazole on the earthworm Aporrectodea icterica.

Earthworms play a key role in agroecosystem soil processes. This study aims to assess the effects of different doses of a commercial formulation of epoxiconazole (Opus®), a persistent and widely used fungicide, on the earthworm Aporrectodea icterica. A laboratory study was conducted in a natural soil in order to measure effects of Opus® on earthworm mortality, uptake, weight gain, enzymatic activities (catalase and glutathione-S-transferase), and energy resources (lipids and glycogens). The estimated LC50 was 45.5 mg kg(-1), or 268 times the recommended dose. Weight gains were 28, 19, and 13% of the initial weight after 28 days of exposure in the control and D1 and D10 (1 and 10 times the recommended dose) treatments, respectively. No difference was observed for catalase activity between the three treatments, at 7, 14, or 28 days. The glutathion-S-transferase (GST) activity was two times as high in D1 as in D0 at 14 days. At 28 days, glycogen concentration was lower in D10 than in the D1 treatment. This study highlighted moderate sublethal effects of the commercial formulation Opus® for earthworms. Considering that these effects were observed on a species found in cultivated fields, even at recommended rates, much more attention should be paid to this pesticide.

The transmembrane protein bacterioopsin affects the polarity of the hydrophobic core of the host lipid bilayer.

Influence of the transmembrane protein bacterioopsin (the retinal-free form of bacteriorhodopsin) on the polarity of egg-phosphatidylcholine bilayers was studied by means of a steady-state and time-resolved fluorescence approach exploiting the solvatochromic properties of the 2-anthroyl fluorophore. Introduced in phosphatidylcholine molecules in the form of 8-(2-anthroyl)octanoic acid, this fluorophore probed the hydrocarbon core of the lipid bilayer. As previously shown (E. Pérochon et al., Biochemistry 31 (1992) 7672-7682), water molecules were detected in this region of the terminal part of the lipid acyl chains. Their number was considerably reduced upon addition of bacterioopsin to the lipids. This was assessed by a blue shift in the fluorescence emission spectra of the probe and a marked decrease in the fractional population of fluorophores interacting with water, to the benefit of those experiencing a hydrophobic environment. In agreement with current theories, this decrease in the hydration of the bilayer may be linked to an increase in the acyl chain order and a decrease in the lateral diffusion coefficient of lipids near the protein. The data obtained at high protein concentration accounts for a protein/lipid interface which is much less hydrated than the hydrophobic core of a protein-free lipid bilayer.

The biotrophic, non-appressorium-forming grass pathogen Claviceps purpurea needs a Fus3/Pmk1 homologous mitogen-activated protein kinase for colonization of rye ovarian tissue.

Claviceps purpurea is a common pathogen of a wide range of grasses and cereals that is able to establish a stable, balanced interaction with its host plant and is considered a biotroph. It does not form special penetration structures such as appressoria. To study the signaling processes involved in this special host-pathogen interaction, we have cloned a gene, cpmk1, encoding a mitogen-activated protein (MAP) kinase that shows significant homology to Fus3 of Saccharomyces cerevisiae and to pmk1 of Magnaporthe grisea. Using a gene-replacement approach, we isolated a Acpmk1 mutant and characterized it in detail. Loss of CPMK1 has no obvious effect on vegetative properties (such as growth rate, morphology, and conidia formation); however, infection tests on rye show that the mutant is unable to colonize rye tissue, i.e., it appears to be completely nonpathogenic. Complementation of the mutant with a wild-type copy of cpmk1 fully restores its pathogenicity, confirming that this MAP kinase is essential for infection of rye by C. purpurea. Transformation of the delta pmk1 mutant of M. grisea with a complete copy of cpmk1 (including the C. purpurea promoter) fully restored its ability to form appressoria and its pathogenicity on barley. Although both fungi drastically differ in their pathogenic strategies, this result indicates that the signal pathway involving CPMK1 is highly conserved.

Gain of virulence caused by insertion of a Pot3 transposon in a Magnaporthe grisea avirulence gene.

The avirulence gene AVR-Pita in Magnaporthe grisea prevents the fungus from infecting rice cultivars carrying the disease resistance gene Pi-ta. Insertion of Pot3 transposon into the promoter of AVR-Pita caused the gain of virulence toward Yashiro-mochi, a rice cultivar containing Pi-ta, which demonstrated the ability of Pot3 to move within the M. grisea genome. The appearance of Pot3 in M. grisea seems to predate the diversification of various host-specific forms of the fungus.

Mapping of avirulence genes in the rice blast fungus, Magnaporthe grisea, with RFLP and RAPD markers.

Three genetically independent avirulence genes, AVR1-Irat7, AVRI-MedNoi; and AVR1-Ku86, were identified in a cross involving isolates Guy11 and 2/0/3 of the rice blast fungus, Magnaporthe grisea. Using 76 random progeny, we constructed a partial genetic map with restriction fragment length polymorphism (RFLP) markers revealed by probes such as the repeated sequences MGL/MGR583 and Pot3/MGR586, cosmids from the M. grisea genetic map, and a telomere sequence oligonucleotide. Avirulence genes AVR1-MedNoi and AVR1-Ku86 were closely linked to telomere RFLPs such as marker TelG (6 cM from AVR1-MedNoi) and TelF (4.5 cM from AVR1-Ku86). Avirulence gene AVR1-Irat7 was linked to a cosmid RFLP located on chromosome 1 and mapped at 20 cM from the avirulence gene AVR1-CO39. Using bulked segregant analysis, we identified 11 random amplified polymorphic DNA (RAPD) markers closely linked (0 to 10 cM) to the avirulence genes segregating in this cross. Most of these RAPD markers corresponded to junction fragments between known or new transposons and a single-copy sequence. Such junctions or the whole sequences of single-copy RAPD markers were frequently absent in one parental isolate. Single-copy sequences from RAPD markers tightly linked to avirulence genes will be used for positional cloning.

Transposon impala, a novel tool for gene tagging in the rice blast fungus Magnaporthe grisea.

impala, a Tc1-mariner transposable element from Fusarium oxysporum, was introduced into the rice blast fungus Magnaporthe grisea to develop transposon-based insertional mutagenesis. A construct (pNIL160) containing an autonomous impala copy inserted in the promoter of niaD encoding Aspergillus nidulans nitrate reductase was introduced by transformation into a M. grisea nitrate reductase-deficient mutant. impala excision was monitored by restoration of prototrophy for nitrate. Southern analysis of niaD+ revertants revealed that impala was able to excise and reinsert at new loci in M. grisea. As observed for its host Fusarium oxysporum, impala inserted at a TA site left a typical excision footprint of 5 bp. We have shown that a defective impala copy was inactive in M. grisea, yet it can be activated by a functional impala transposase. A transformant carrying a single copy of pNIL160 was used to generate a collection of 350 revertants. Mutants either altered for their mycelial growth (Rev2) or nonpathogenic (Rev77) were obtained. Complementation of Rev77 with a 3-kb genomic fragment from a wild-type locus was successful, demonstrating the tagging of a pathogenicity gene by impala. This gene, called ORP1, is essential for penetration of host leaves by M. grisea and has no sequence homology to known genes.

Expression cloning in Fe2+ transport defective yeast of a novel maize MYC transcription factor.

A complementation approach of the yeast fet3fet4 mutant strain, defective in both low- and high-affinity iron transport, was initiated as an attempt to characterize the Fe(III)-mugineic acid (MA) transporter from grasses. A maize cDNA encoding a novel MYC transcription factor, named 7E, was cloned by screening an iron-deficient maize root cDNA expression library on a minimum media containing Fe(III)-deoxyMA as a unique iron source. 7E expression restored growth specifically to the fet3 fet4 mutant strain. It did not affect growth rate of a trk1trk2 potassium transport defective yeast strain or parental W303 strain growth rate. No 55Fe uptake increase was observed in 7E transformed fet3 fet4 yeast during short-term kinetics. However, the iron accumulation in these cells was 1.3-fold higher than in untransformed cells after a 24-h period. The 7E protein contained 694 amino acids and had a predicted molecular mass of 74.2kDa. It had 44% identity with the RAP-1 protein, a 67.9-kDa MYC-like protein from Arabidopsis thaliana which binds the G-box sequence via a basic region helix-loop-helix (bHLH), without requiring heterodimerization with MYB proteins. Phylogenic comparisons revealed that the maize 7E protein was related to the Arabidopsis thaliana RAP-1 protein and to the Phaseolus vulgaris PG1. This similarity was particularly evident for the bHLH domain, which was 95% identical between maize 7E and Arabidopsis thaliana RAP-1. 7E, RAP-1 and PG-1 proteins revealed a plant MYC-like sub-family that was more related to the maize repressor-like IN1 than to maize R proteins. 7E mRNA was detected in both roots and leaves by the Northern analysis. The amount of 7E mRNA increased, in response to iron starvation, by 20 and 40% in roots and leaves, respectively. The relationship between iron metabolism and myc expression in animal cells is discussed.

Is the protein/lipid hydrophobic matching principle relevant to membrane organization and functions?

Biological membranes are complex and well-organized multimolecular assemblies composed of a wide variety of protein and lipid molecular species. If such a diversity in protein and lipid polar headgroup structures may easily be related to a large panel of functions, the wide dispersion in acyl chain length and structure which the lipids display is more difficult to understand. It is not required for maintaining bilayer assembly and fluidity. Direct information on the lateral distribution of these various molecular species, on their potential specificity for interaction between themselves and with proteins and on the functional implications of these interactions is also still lacking. Because hydrophobic interactions play a major role in stabilizing membrane structures, we suggest considering the problem from the point of view of the matching of the hydrophobic surface of proteins by the acyl chains of the lipids. After a brief introduction to the hydrophobic matching principle, we will present experimental results which demonstrate the predictive power of the current theories and then, we will introduce the new and important concept of protein/lipid sorting in membranes. Finally, we will show how the hydrophobic matching condition may play a key role in the membrane organization and function.