FUM1--a gene required for fumonisin biosynthesis but not for maize ear rot and ear infection by Gibberella moniliformis in field tests

We have analyzed the role of fumonisins in infection of maize (Zea mays) by Gibberella moniliformis (anamorph Fusarium verticillioides) in field tests in Illinois and Iowa, United States. Fumonisin-nonproducing mutants were obtained by disrupting FUM1 (previously FUM5), the gene encoding a polyketide synthase required for fumonisin biosynthesis. Maize ear rot, ear infection, and fumonisin contamination were assessed by silk-channel injection in 1999 and 2000 and also by spray application onto maize silks, injection into maize stalks, and application with maize seeds at planting in 1999. Ear rot was evaluated by visual assessment of whole ears and by calculating percentage of symptomatic kernels by weight. Fumonisin levels in kernels were determined by high-performance liquid chromatography. The presence of applied strains in kernels was determined by analysis of recovered isolates for genetic markers and fumonisin production. Two independent fumonisin-nonproducing (fum1-3 and fum1-4) mutants were similar to their respective fumonisin-producing (FUM1-1) progenitor strains in ability to cause ear rot following silk-channel injection and also were similar in ability to infect maize ears following application by all four methods tested. This evidence confirms that fumonisins are not required for G. moniliformis to cause maize ear rot and ear infection.     

Leishmania promastigotes require lipophosphoglycan to actively modulate the fusion properties of phagosomes at an early step of phagocytosis

The lipophosphoglycan (LPG) of Leishmania promastigotes plays key roles in parasite survival in both insect and mammalian hosts. Evidence suggests that LPG decreases phagosome fusion properties at the onset of infection in macrophages. The mechanisms of action of this molecule are, however, poorly understood. In the present study, we used a panoply of Leishmania mutants displaying modified LPG structures to determine more precisely how LPG modulates phagosome-endosome fusion. Using an in vivo fusion assay measuring, at the electron microscope, the transfer of solute materials from endosomes to phagosomes, we provided further evidence that the repeating Gal(beta1,4)Man(alpha1-PO4) units of LPG are responsible for the alteration in phagosome fusion. The inhibitory effect of LPG on phagosome fusion was shown to be more potent towards late endocytic organelles and lysosomes than early endosomes, explaining how Leishmania promastigotes can avoid degradation in hydrolase-enriched compartments. The involvement of other repeating unit-containing molecules, including the secreted acid phosphatase, in the inhibition process was ruled out, as an LPG-defective mutant (Ipg1-) which secretes repeating unit-containing glycoconjugates was present in highly fusogenic phagosomes. In L. major, oligosaccharide side-chains of LPG did not contribute to the inhibition process, as Spock, an L. major mutant lacking LPG side-chains, blocked fusion to the same extent as wild-type parasites. Finally, dead parasites internalized from the culture medium were not as efficient as live parasites in altering phagosome-endosome fusion, despite the presence of LPG. However, the killing of parasites with vital dyes after their sequestration in phagosomes had no effect on the fusion properties of this organelle. Collectively, these results suggest that living promastigotes displaying full-length cell surface LPG can actively influence macrophages at an early stage of phagocytosis to generate phagosomes with poor fusogenic properties.     

Masculinized dominant females in a cooperatively breeding species

The molecular mechanisms underlying complex social behaviours such as dominance are largely unknown. Studying the cooperatively breeding African cichlid Neolamprologus pulcher, we show that dominant females were similar to dominant males in dominance behaviour, high testosterone levels and brain arginine vasotocin expression (a neuropeptide involved in vertebrate territorial, reproductive and social behaviours) compared to subordinate helpers, but had lower levels of 11-ketotestosterone than males. Furthermore, brain gene expression profiles of dominant females were most similar to those of the males (independent of social rank). Dominant breeder females are masculinized at the molecular and hormonal level while being at the same time reproductively competent, suggesting a modular organization of molecular and endocrine functions, allowing for sex-specific regulation.     

RUN and FYVE domain–containing protein 4 enhances autophagy and lysosome tethering in response to Interleukin-4

Autophagy is a key degradative pathway coordinated by external cues, including starvation, oxidative stress, or pathogen detection. Rare are the molecules known to contribute mechanistically to the regulation of autophagy and expressed specifically in particular environmental contexts or in distinct cell types. Here, we unravel the role of RUN and FYVE domain–containing protein 4 (RUFY4) as a positive molecular regulator of macroautophagy in primary dendritic cells (DCs). We show that exposure to interleukin-4 (IL-4) during DC differentiation enhances autophagy flux through mTORC1 regulation and RUFY4 induction, which in turn actively promote LC3 degradation, Syntaxin 17–positive autophagosome formation, and lysosome tethering. Enhanced autophagy boosts endogenous antigen presentation by MHC II and allows host control of Brucella abortus replication in IL-4–treated DCs and in RUFY4-expressing cells. RUFY4 is therefore the first molecule characterized to date that promotes autophagy and influences endosome dynamics in a subset of immune cells.     

Multiplex PCR/liquid chromatography assay for detection of gene rearrangements: application to RB1 gene

Screening for large gene rearrangements is established as an important part of molecular medicine but is also challenging. A variety of robust methods can detect whole-gene deletions, but will fail to detect more subtle rearrangements that may involve a single exon. In this paper, we describe a new, versatile and robust method to assess exon copy number, called multiplex PCR/liquid chromatography assay (MP/LC). Multiple exons are amplified using unlabeled primers, then separated by ion-pair reversed-phase high-performance liquid chromatography (IP-RP-HPLC), and quantitated by fluorescent detection using a post-column intercalation dye. The relative peak intensities for each target directly reflect exon copy number. This novel technique was used to screen a panel of 121 unrelated retinoblastoma patients who were tested previously using a reference strategy. MP/LC correctly scored all deletions and demonstrated a previously undetected RB1 duplication, the first to be described. MP/LC appears to be an easy, versatile, and cost-effective method, which is particularly relevant to denaturing HPLC (DHPLC) users since it broadens the spectrum of available applications on a DHPLC system.     

Evidence for size and sex-specific dispersal in a cooperatively breeding cichlid fish

African Great Lake cichlid populations are divided into thousands of genetic subpopulations. The low gene flow between these subpopulations is thought to result from high degrees of natal philopatry, heavy predation pressure, and a patchy distribution of preferred habitats. While predation pressure and habitat distribution are fairly straightforward to assess, data on dispersal distances and rates are scarce. In fishes, direct observations of dispersal events are unlikely, but dispersal can be studied using molecular markers. Using seven microsatellite loci, we examined dispersal in the cooperatively breeding cichlid fish, Neolamprologus pulcher. As this species is found in well-defined groups clustered into subpopulations, we could assess dispersal on a narrow (within subpopulation) and broad (between subpopulation) scale. While fish were generally more related to others in their own subpopulation than they were to fish from other subpopulations, large males diverged from this pattern. Large males were more related to other large males from different subpopulations than they were to large males from their own subpopulation, suggesting more frequent dispersal by large males. Across subpopulations, relatedness between large males was higher than the relatedness among large females; this pattern was not detected in small males and small females. Within a subpopulation, individuals appeared to be preferentially moving away from relatives, and movement was unrestricted by the physical distance between groups. Our results highlight the importance of examining multiple spatial scales when studying individual dispersal biases.     

Biomechanical assessment of sitting pivot transfer tasks using a newly developed instrumented transfer system among long-term wheelchair users

This paper describes the technical characteristics of a transfer assessment system, along with details on three-dimensional (3D) upper extremity (U/E) kinematics required to compute U/E joint forces and moments using inverse dynamics during a displacement of the body in a sitting position from an initial surface to a target one (sitting pivot transfer (SPT)). This system includes five instrumented surfaces designed to measure position (center of pressure (COP)), magnitude and direction of the tri-axial force components underneath the feet, hands (leading and trailing) and buttocks (initial and target seats) during SPTs. Linearity, COP position and natural frequency tests were performed to confirm the accuracy of the transfer assessment system outcomes. Preliminary data of one person with spinal cord injury performing SPTs toward a target seat of same height (50 cm) and additional ones toward a raised target seat (60 cm) are presented. The transfer assessment system was found to be safe, versatile in terms of height- and width-adjustment ranges, portable within a laboratory environment, easy for experienced rehabilitation scientists to use, and allowed for valid quantification of reaction forces during SPTs as confirmed by the overall accuracy test results. Combined with the 3D U/E kinematic and anthropometric parameters, the transfer assessment system outcomes allowed for the quantification of U/E joint forces and moments. Preliminary results highlight the kinematic and kinetic specificities of the leading and trailing shoulders and elbows during SPTs. The impact of modifying target seat heights on the kinematic and kinetic outcomes during SPTs is explored. The transfer assessment framework proposed is useful for research and offers a wide spectrum of possibilities for acquiring new biomechanical knowledge on SPTs that may strengthen clinical practice guidelines, targeting the preservation of U/E integrity following SCI.     

Cryptic speciation in Fusarium subglutinans

Fusarium isolates that form part of the Gibberella fujikuroi species complex have been classified using either a morphological, biological, or phylogenetic species concept. Problems with the taxonomy of Fusarium species in this complex are mostly experienced when the morphological and biological species concepts are applied. The most consistent identifications are obtained with the phylogenetic species concept. Results from recent studies have presented an example of discordance between the biological and phylogenetic species concepts, where a group of F. subglutinans sensu stricto isolates, i.e., isolates belonging to mating population E of the G. fujikuroi complex, could be sub-divided into more than one phylogenetic lineage. The aim of this study was to determine whether this sub-division represented species divergence or intraspecific diversity in F. subglutinans. For this purpose, we included 29 F. subglutinans isolates belonging to the E-mating population that were collected from either maize or teosinte, from a wide geographic range. DNA sequence data for six nuclear regions in each of these isolates were obtained and used in phylogenetic concordance analyses. These analyses revealed the presence of two major groups representing cryptic species in F. subglutinans. These cryptic species were further sub-divided into a number of smaller groups that appear to be reproductively isolated in nature. This suggests not only that the existing F. subglutinans populations are in the process of divergence, but also that each of the resulting lineages are undergoing separation into distinct taxa. These divergences did not appear to be linked to geographic origin, host, or phenotypic characters such as morphology.     

Ethylene triggers needle abscission in root-detached balsam fir

Post-harvest needle abscission is a major challenge for Christmas tree and greenery industries. It was hypothesized that ethylene triggers abscission in balsam fir. Three experiments were conducted to test this hypothesis. In experiment 1, 70 balsam fir branches were collected, placed in water, and ethylene evolution was observed over time. In experiment 2, a 2 × 5 factorial experiment was designed to determine the effect of exogenous ethylene and an ethylene receptor blocker, 1-methylcyclopropene (1-MCP), on needle abscission. In experiment 3, a 2 × 6 factorial experiment was designed to determine the effect of exogenous ethylene and an ethylene inhibitor, aminoethoxyvinylglycine (AVG), on needle abscission. It was found that ethylene evolution was the highest 1–2 days prior to needle abscission, which was consistent in untreated branches and branches exposed to exogenous ethylene. Exposure to exogenous ethylene significantly decreased needle retention by 63%. When ethylene receptors were blocked by 1-MCP, needle retention increased by 147% despite the presence of ethylene and increased by 73% in the absence of ethylene when compared to the respective controls. When endogenous ethylene synthesis was inhibited by AVG, there was no improvement in needle retention in the presence of ethylene, but there was a 113% increase in needle retention in the absence of exogenous ethylene. Ethylene is strongly implicated as the signal triggering abscission in root-detached balsam fir.