C9orf72 ALS/FTD dipeptide repeat protein levels are reduced by small molecules that inhibit PKA or enhance protein degradation

Intronic GGGGCC (G4C2) hexanucleotide repeat expansion within the human C9orf72 gene represents the most common cause of familial forms of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) (C9ALS/FTD). Repeat-associated non-AUG (RAN) translation of repeat-containing C9orf72 RNA results in the production of neurotoxic dipeptide-repeat proteins (DPRs). Here, we developed a high-throughput drug screen for the identification of positive and negative modulators of DPR levels. We found that HSP90 inhibitor geldanamycin and aldosterone antagonist spironolactone reduced DPR levels by promoting protein degradation via the proteasome and autophagy pathways respectively. Surprisingly, cAMP-elevating compounds boosting protein kinase A (PKA) activity increased DPR levels. Inhibition of PKA activity, by both pharmacological and genetic approaches, reduced DPR levels in cells and rescued pathological phenotypes in a Drosophila model of C9ALS/FTD. Moreover, knockdown of PKA-catalytic subunits correlated with reduced translation efficiency of DPRs, while the PKA inhibitor H89 reduced endogenous DPR levels in C9ALS/FTD patient-derived iPSC motor neurons. Together, our results suggest new and druggable pathways modulating DPR levels in C9ALS/FTD

Planck pre-launch status : The Planck mission

Peer reviewe

Temperature and climate affect the endophytes community in grapevine

Science has just started understanding how the environment drives the composition of microbial communities. Endophytes, as host-associated microbes, respond to environmental stimuli in a host-mediated fashion. To study how temperature and climate may affect endophytic microbial communities we studied grapevine-associated microbial populations using a cultivation independent approach. Grafted cuttings were used to analyse how temperature affects microbial endophytic communities in a controlled environment. The composition of microbial endophytes in the field was assessed by surveying potted plants at different altitudes representing diverse climatic conditions. Seasonal fluctuations in the microbial endophytes were also considered by sampling test plants at different times throughout the year. We adopted a DNA-based, cultivation-independent approach to the analysis of microbial populations variability in the conditions considered in this study. The analysis of DNA amplified by PCR involved the use of both Automated Ribosomal Intergenic Spacer Analysis (ARISA) and Roche 454 GS FLX+ technology

Metagenomic analysis of bacterial endophytic communities associated with grapevine (Vitis vinifera L.)

In recent years, interest in endophytic microorganisms has increased, as they play a key role in agricultural environments and are promising because of their potential use in sustainable agriculture. These microorganisms include both commensal species, which have no direct effect on the host plant, and mutualistic symbionts, which could be used in the biological control of pathogens or plant growth promotion. In the present study we investigated how microbial communities in plants from organically managed farms differ from those obtained from integrated pest management (IPM) farms. Microbial DNA isolated from grapevines (Vitis vinifera L.) cv Merlot and Chardonnay cultivated in a subalpine area in Northern Italy was PCR amplified to fingerprint endophytic communities, and to assess the distribution of important functional genes in the grapevine microbiome in the studied areas. Here we report the composition of endophytic microbial communities assessed through a cultivation independent approach: Automated Ribosomal Intergenic Spacer Analysis (ARISA). The changes in community structure and composition are interpreted in the light of the environmental variables considered. Fingerprinting results were validated by multivariate analysis. Other metagenomics approaches are being considered

How grapevine got pimples: the interkingdom horizontal transfer of an unusual symbiont

Humans have established a long-lasting co-habitation with a variety of animals, plants and microbes. The human-gut microbe interaction has been recently explored, and cases have been reported of microbial host transfer from domesticated animals to humans. Much less is known regarding human-plant microbial transfers. Here we report a recently established symbiosis between the human-associated opportunistic pathogen Propionibacterium acnes and the highly domesticated grape Vitis vinifera. We detected P. acnes in many grape plants using pyrosequencing of 16S rDNA and fluorescent in-situ hybridization (FISH) which localized P. acnes in specific endosphere habitats (pith and bark). We also show that recA genes in uncultivable endophytic P. acnes of grape are pseudogenes, suggesting a recent obligate symbiosis which we tentative date as being not older than 7000 years ago, an age compatible with the domestication of grape by humans. Our results represent the first documented inter-kingdom horizontal host transfer of a human symbion

Characterisation of microbial endophytes from wild grapevine (Vitis vinifera subsp. sylvestris)

While a large number of studies aim at analyzing the microorganisms found in crops, using either a classical microbiology approach or more novel, DNA-based techniques, little is known on the selective forces affecting these microbial communities during the process of domestication or if and how they shaped plant-associated microbes. We are interested in elucidating how domestication has influenced bacteria and fungi colonizing the grapevine endosphere, and how these modifications interfere with plant physiology, growth and health. We isolated microbial endophytes from V. vinifera subsp. sylvestris plants obtained from different areas of Italy and analysed the isolates for important traits related to interaction with the plant host, social behaviour, tolerance to antibiotics and their production, biocontrol. In addition, a DNA-dependent approach was adopted to fingerprint microbial communities in wild grapevine plants and to compare theme to non-domesticated plant. Automated Ribosomal Intergenic Spacer Analysis (ARISA) was used to assess variability and identity of the non culturable microbial fauna. Both geographical origin and plant genotype were considered. The ability of some of these isolates to colonize domesticated grapevines was investigated to assess their viability for reintroduction in cultivated grapevines

Disease management in grapevine and fungal endophytic populations

Grapevine tissues are colonised by a variety of endophytic fungi with no detectable involvement in pathogenesis. The roles of the non pathogenic mycota associated with internal tissues are still not fully understood. Endophytic fungi are often considered an (as yet) untapped source of biocontrol microorganisms, and a reservoir of biological functions plants can benefit from. More recently, a role for some endophytic species as potential allies of incoming pathogens was hypothesised. As grapevines are intensively subjected to the application of antifungal compounds, we investigated the impact of different agricultural practices on endophytic fungi. The analysis of distribution of culturable fungi was integrated by a DNA-dependant approach, involving fingerprinting the ITS regions of all strains colonising the studied plants. Plants from cultivars Chardonnay and Merlot were sampled, from both organic and IPM vineyards, in seven different areas of a renowned Italian wine-growing county (Trentino). For comparison, endophytic fungi and total DNA were isolated from stems of Vitis vinifera subsp. sylvestris plants. We found the composition of fungal endophytic communities in plants of cultivated grapevines (V. vinifera subsp. vinifera) to be highly homogeneous. The prevalent taxa were Alternaria spp., Epicoccum nigrum and Aureobasidium pullulans. Multivariate statistical analysis showed that fungal communities were different between plants form vineyards adopting organic and IPM management, supporting the hypothesis that vineyard management affects non pathogenic fungi associated with plant tissues. A slighter difference was observed between fungi associated with grapevines of different cultivars. A largely different microbial community was detected on wild grapevines (V. vinifera subsp. sylvestris). The prevalent fungal isolates identified from the tissues of wild grapevine were Cladosporium sp., Acremonium sp., Cryptococcus sp. and to a lesser extent, Alternaria sp. The ability of new fungal endophytes, isolated from wild grapevine, to colonise domesticated grapevines is being studied to understand if these new tools can be applied for plant biocontrol purposes

Profiling of grapevine fungal endophytic community using Automated Ribosomal Intergenic Spacer Analysis (ARISA)

A wide diversity of endophytic fungi has been associated with grapevines (Vitis vinifera L.). The composition of the endophytic microbial population changes greatly across plants and plant parts. The factors affecting the fluctuations of fungal communities composition in grapevine are as yet not understood. We compared fungal endophytic populations in Italian grapevines across farms using organic or integrated pest management (IPM). Both isolation of fungi on synthetic media and community analysis using an automated, DNA-based approach (ARISA) agreed in indicating that fungal populations from organic farms were significantly different from those from IPM farms. A multivariate statistical analysis showed that data obtained from DNA-based community fingerprinting ARISA was more information-rich than the analysis of cultured fungi