Tum—mutation P35B generates the MHC-binding site of a new antigenic peptide

Immunogenic tumor cell variant P35 was obtained by mutagen treatment of mouse mastocytoma P815. It express a potent new antigen recognized by syngeneic cytolytic T lymphocytes (CTL). This antigen is the result of a point mutation in a gene that is expressed by most healthy cells. A decapeptide encoded by the region spanning the mutation sensitized P815 cells to the relevant CTL, whereas the homologous decapeptide corresponding to the normal sequence did not. Only the mutant decapeptide was capable of enhancing the expression of the D^d-presenting molecule at the cell surface, indicating that the mutation generates a motif which enables the antigenic peptide to bind to D^d. Correspondence to: T. Boon.     

Non-histone protein methylation in Trypanosoma cruzi epimastigotes

Post-translational methylation of proteins, which occurs in arginines and lysines, modulates several biological processes at different levels of cell signaling. Recently, methylation has been demonstrated in the regulation beyond histones, for example, in the dynamics of protein-protein and protein-nucleic acid interactions. However, the presence and role of non-histone methylation in Trypanosoma cruzi, the etiologic agent of Chagas disease, has not yet been elucidated. Here, we applied mass spectrometry-based-proteomics (LC-MS/MS) to profile the methylproteome of T. cruzi epimastigotes, describing a total of 1252 methyl sites in 824 proteins. Functional enrichment and protein-protein interaction analysis show that protein methylation impacts important biological processes of the parasite, such as translation, RNA and DNA binding, amino acid, and carbohydrate metabolism. In addition, 171 of the methylated proteins were previously reported to bear phosphorylation sites in T. cruzi, including flagellar proteins and RNA binding proteins, indicating that there may be an interplay between these different modifications in non-histone proteins. Our results show that a broad spectrum of functions is affected by methylation in T. cruzi, indicating its potential to impact important processes in the biology of the parasite and other trypanosomes.     

Expansion microscopy allows high resolution single cell analysis of epigenetic readers

Interactions between epigenetic readers and histone modifications play a pivotal role in gene expression regulation and aberrations can enact etiopathogenic roles in both developmental and acquired disorders like cancer. Typically, epigenetic interactions are studied by mass spectrometry or chromatin immunoprecipitation sequencing. However, in these methods, spatial information is completely lost. Here, we devise an expansion microscopy based method, termed Expansion Microscopy for Epigenetics or ExEpi, to preserve spatial information and improve resolution. We calculated relative co-localization ratios for two epigenetic readers, lens epithelium derived growth factor (LEDGF) and bromodomain containing protein 4 (BRD4), with marks for heterochromatin (H3K9me3 and H3K27me3) and euchromatin (H3K36me2, H3K36me3 and H3K9/14ac). ExEpi confirmed their preferred epigenetic interactions, showing co-localization for LEDGF with H3K36me3/me2 and for BRD4 with H3K9/14ac. Moreover addition of JQ1, a known BET-inhibitor, abolished BRD4 interaction with H3K9/14ac with an IC₅₀ of 137 nM, indicating ExEpi could serve as a platform for epigenetic drug discovery. Since ExEpi retains spatial information, the nuclear localization of marks and readers was determined, which is one of the main advantages of ExEpi. The heterochromatin mark, H3K9me3, is located in the nuclear rim whereas LEDGF co-localization with H3K36me3 and BRD4 co-localization with H3K9/14ac occur further inside the nucleus.     

Interaction of endophytic diazotrophic bacteria and Fusarium oxysporum f. sp. cubense on plantlets of banana ‘Maça’

The objective of this work was to evaluate the effect of endophytic diazotrophic bacteria Herbaspirillum and Burkholderia on the Fusarium oxysporum f. sp. cubense (Foc) establishment and on the plantlets growth of the ‘Maçã’ banana (Musa spp., group ABB). Two assays were carried out in a greenhouse at the National Center of Tropical Agroindustry in Fortaleza city, Ceará State (Brazil), using randomized block designs in the factorial arrangements 4?×?2 and 8?×?2. On the first trial plantlets were inoculated with the Burkholderia sp. AB202; Herbaspirillum sp., BA227, both of strains and controls bacteria, with and without Foc and cultivated in pots filled with an autoclaved mixture of washed sand and vermiculite (ratio 3:2 v v^?1), during 4 months. On a second assay, plants were subjected to following conditions: absence and presence of strains AB202, AB213 (Burkholderia spp.), BA227, BA234 (Herbaspirillum-like), AB202 plus BA227, AB213 plus BA234, the mixture of the four bacterial strain, absence and presence of the Foc; and cultivated in pots filled with autoclaved Haplic Arenosol, during 2 months. The plant association with diazotrophs and the Foc was confirmed, and factors interacted significantly on the most probable number of bacteria and the colony forming units of the pathogen on roots and plant rhizomes. The potential of the endophytes on the inhibition of Foc propagate units and on the plant growth promotion was demonstrated. The higher biomass was observed four and 2 months after plant inoculation with AB202 and BA234. Results showed that these endophytes may be used as potential biofertilizer and biocontrol agents.     

Phylogeography of the Petunia integrifolia complex in southern Brazil

In this study, we evaluated the genetic diversity of the Petunia integrifolia species group using a phylogeographical approach, and attempted to understand better its diversification and taxonomy. Plants from five morphological groups were collected, covering a large part of the geographical distribution of most of the species. Two major clades were found in the phylogenetic tree, and an additional lineage, corresponding to P. inflata, was found in the haplotype network obtained for plastid markers. All three lineages are clearly delimited geographically, but, with the exception of P. inflata, the morphological groups were not genetically distinct. Our results suggest that a population expansion after a size reduction resulted in the establishment of two distinct and allopatric groups c. 0.5 Mya, one group occurring in a geologically ancient area, and the other occurring in areas that were under the influence of a series of marine transgressions/regressions at the end of the Pleistocene. These two clades are evolutionarily significant units with significantly different allele frequencies in their nuclear genome and reciprocal monophyly in maternal, uniparentally inherited markers. All our results suggest that the morphology‐based taxonomy in this group does not reflect its evolutionary history, and revision of its species limits should incorporate the distribution of the genetic diversity. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 174 , 199–213.     

Inventory and monitoring of wine microbial consortia

The evolution of the wine microbial ecosystem is generally restricted to Saccharomyces cerevisiae and Oenococcus oeni, which are the two main agents in the transformation of grape must into wine by acting during alcoholic and malolactic fermentation, respectively. But others species like the yeast Brettanomyces bruxellensis and certain ropy strains of Pediococcus parvulus can spoil the wine. The aim of this study was to address the composition of the system more precisely, identifying other components. The advantages of the polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) approach to wine microbial ecology studies are illustrated by bacteria and yeast species identification and their monitoring at each stage of wine production. After direct DNA extraction, PCR-DGGE was used to make the most exhaustive possible inventory of bacteria and yeast species found in a wine environment. Phylogenetic neighbor-joining trees were built to illustrate microbial diversity. PCR-DGGE was also combined with population enumeration in selective media to monitor microbial changes at all stages of production. Moreover, enrichment media helped to detect the appearance of spoilage species. The genetic diversity of the wine microbial community and its dynamics during winemaking were also described. Most importantly, our study provides a better understanding of the complexity and diversity of the wine microbial consortium at all stages of the winemaking process: on grape berries, in must during fermentation, and in wine during aging. On grapes, 52 different yeast species and 40 bacteria could be identified. The diversity was dramatically reduced during winemaking then during aging. Yeast and lactic acid bacteria were also isolated from very old vintages. B. bruxellensis and O. oeni were the most frequent.     

Plasmodium berghei leucine-rich repeat protein 1 downregulates protein phosphatase 1 activity and is required for efficient oocyst development

Protein phosphatase 1 (PP1) is a key enzyme for Plasmodium development. However, the detailed mechanisms underlying its regulation remain to be deciphered. Here, we report the functional characterization of the Plasmodium berghei leucine-rich repeat protein 1 (PbLRR1), an orthologue of SDS22, one of the most ancient and conserved PP1 interactors. Our study shows that PbLRR1 is expressed during intra-erythrocytic development of the parasite, and up to the zygote stage in mosquitoes. PbLRR1 can be found in complex with PbPP1 in both asexual and sexual stages and inhibits its phosphatase activity. Genetic analysis demonstrates that PbLRR1 depletion adversely affects the development of oocysts. PbLRR1 interactome analysis associated with phospho-proteomics studies identifies several novel putative PbLRR1/PbPP1 partners. Some of these partners have previously been characterized as essential for the parasite sexual development. Interestingly, and for the first time, Inhibitor 3 (I3), a well-known and direct interactant of Plasmodium PP1, was found to be drastically hypophosphorylated in PbLRR1-depleted parasites. These data, along with the detection of I3 with PP1 in the LRR1 interactome, strongly suggest that the phosphorylation status of PbI3 is under the control of the PP1–LRR1 complex and could contribute (in)directly to oocyst development. This study provides new insights into previously unrecognized PbPP1 fine regulation of Plasmodium oocyst development through its interaction with PbLRR1.     

Role of the hindbrain in patterning the otic vesicle: a study of the zebrafish vhnf1 mutant

The vertebrate inner ear develops from an ectodermal placode adjacent to rhombomeres 4 to 6 of the segmented hindbrain. The placode then transforms into a vesicle and becomes regionalised along its anteroposterior, dorsoventral and mediolateral axes. To investigate the role of hindbrain signals in instructing otic vesicle regionalisation, we analysed ear development in zebrafish mutants for vhnf1, a gene expressed in the caudal hindbrain during otic induction and regionalisation. We show that, in vhnf1 homozygous embryos, the patterning of the otic vesicle is affected along both the anteroposterior and dorsoventral axes. First, anterior gene expression domains are either expanded along the whole anteroposterior axis of the vesicle or duplicated in the posterior region. Second, the dorsal domain is severely reduced, and cell groups normally located ventrally are shifted dorsally, sometimes forming a single dorsal patch along the whole AP extent of the otic vesicle. Third, and probably as a consequence, the size and organization of the sensory and neurogenic epithelia are disturbed. These results demonstrate that, in zebrafish, signals from the hindbrain control the patterning of the otic vesicle, not only along the anteroposterior axis, but also, as in amniotes, along the dorsoventral axis. They suggest that, despite the evolution of inner ear structure and function, some of the mechanisms underlying the regionalisation of the otic vesicle in fish and amniotes have been conserved.     

NCAM Mimetic Peptides: Potential Therapeutic Target for Neurological Disorders

The neural cell adhesion molecule (NCAM) plays a pivotal role in the development and maintenance of the nervous system via homophilic (NCAM–NCAM) and heterophilic (NCAM-other molecules) interactions. Many synthetic peptides have been engineered to mimic these interactions and induce NCAM-downstream signaling pathways. Such NCAM mimetics have displayed neuritogenic and neuroprotective properties, as well as synaptic modulation in vitro and in vivo. Furthermore, they have been used successfully in preclinical studies to treat neurological disorders including stroke, traumatic brain injury and Alzheimer’s disease. This review focuses on recent progress in the development of NCAM mimetic peptides, in particular, on establishing C3, plannexin, and FGL as therapeutic candidates for neurological disorders.