Lactobacillus delbrueckii ssp. lactis and ssp. bulgaricus: a chronicle of evolution in action

Lactobacillus delbrueckii ssp. lactis and ssp. bulgaricus are lactic acid producing bacteria that are largely used in dairy industries, notably in cheese-making and yogurt production. An earlier in-depth study of the first completely sequenced ssp. bulgaricus genome revealed the characteristics of a genome in an active phase of rapid evolution, in what appears to be an adaptation to the milk environment. Here we examine for the first time if the same conclusions apply to the ssp. lactis, and discuss intra- and inter-subspecies genomic diversity in the context of evolutionary adaptation. RESULTS: Both L. delbrueckii ssp. show the signs of reductive evolution through the elimination of superfluous genes, thereby limiting their carbohydrate metabolic capacities and amino acid biosynthesis potential. In the ssp. lactis this reductive evolution has gone less far than in the ssp. bulgaricus. Consequently, the ssp. lactis retained more extended carbohydrate metabolizing capabilities than the ssp. bulgaricus but, due to high intra-subspecies diversity, very few carbohydrate substrates, if any, allow a reliable distinction of the two ssp.. We further show that one of the most important traits, lactose fermentation, of one of the economically most important dairy bacteria, L. delbruecki ssp. bulgaricus, relies on horizontally acquired rather than deep ancestral genes. In this sense this bacterium may thus be regarded as a natural GMO avant la lettre. CONCLUSIONS: The dairy lactic acid producing bacteria L. delbrueckii ssp. lactis and ssp. bulgaricus appear to represent different points on the same evolutionary track of adaptation to the milk environment through the loss of superfluous functions and the acquisition of functions that allow an optimized utilization of milk resources, where the ssp. bulgaricus has progressed further away from the common ancestor

PURA-Related Developmental and Epileptic Encephalopathy Phenotypic and Genotypic Spectrum

Background and Objectives Purine-rich element-binding protein A (PURA) gene encodes Pur-α, a conserved protein essential for normal postnatal brain development. Recently, a PURA syndrome characterized by intellectual disability, hypotonia, epilepsy, and dysmorphic features was suggested. The aim of this study was to define and expand the phenotypic spectrum of PURA syndrome by collecting data, including EEG, from a large cohort of affected patients. Methods Data on unpublished and published cases were collected through the PURA Syndrome Foundation and the literature. Data on clinical, genetic, neuroimaging, and neurophysiologic features were obtained. Results A cohort of 142 patients was included. Characteristics of the PURA syndrome included neonatal hypotonia, feeding difficulties, and respiratory distress. Sixty percent of the patients developed epilepsy with myoclonic, generalized tonic-clonic, focal seizures, and/or epileptic spasms. EEG showed generalized, multifocal, or focal epileptic abnormalities. Lennox-Gastaut was the most common epilepsy syndrome. Drug refractoriness was common: 33.3% achieved seizure freedom. We found 97 pathogenic variants in PURA without any clear genotype-phenotype associations. Discussion The PURA syndrome presents with a developmental and epileptic encephalopathy with characteristics recognizable from neonatal age, which should prompt genetic screening. Sixty percent have drug-resistant epilepsy with focal or generalized seizures. We collected more than 90 pathogenic variants without observing overt genotype-phenotype associations

Tau-Mediated Nuclear Depletion and Cytoplasmic Accumulation of SFPQ in Alzheimer's and Pick's Disease

Tau dysfunction characterizes neurodegenerative diseases such as Alzheimer's disease (AD) and frontotemporal lobar degeneration (FTLD). Here, we performed an unbiased SAGE (serial analysis of gene expression) of differentially expressed mRNAs in the amygdala of transgenic pR5 mice that express human tau carrying the P301L mutation previously identified in familial cases of FTLD. SAGE identified 29 deregulated transcripts including Sfpq that encodes a nuclear factor implicated in the splicing and regulation of gene expression. To assess the relevance for human disease we analyzed brains from AD, Pick's disease (PiD, a form of FTLD), and control cases. Strikingly, in AD and PiD, both dementias with a tau pathology, affected brain areas showed a virtually complete nuclear depletion of SFPQ in both neurons and astrocytes, along with cytoplasmic accumulation. Accordingly, neurons harboring either AD tangles or Pick bodies were also depleted of SFPQ. Immunoblot analysis of human entorhinal cortex samples revealed reduced SFPQ levels with advanced Braak stages suggesting that the SFPQ pathology may progress together with the tau pathology in AD. To determine a causal role for tau, we stably expressed both wild-type and P301L human tau in human SH-SY5Y neuroblastoma cells, an established cell culture model of tau pathology. The cells were differentiated by two independent methods, mitomycin C-mediated cell cycle arrest or neuronal differentiation with retinoic acid. Confocal microscopy revealed that SFPQ was confined to nuclei in non-transfected wild-type cells, whereas in wild-type and P301L tau over-expressing cells, irrespective of the differentiation method, it formed aggregates in the cytoplasm, suggesting that pathogenic tau drives SFPQ pathology in post-mitotic cells. Our findings add SFPQ to a growing list of transcription factors with an altered nucleo-cytoplasmic distribution under neurodegenerative conditions

Intronic ATTTC repeat expansions in STARD7 in familial adult myoclonic epilepsy linked to chromosome 2

Familial Adult Myoclonic Epilepsy (FAME) is characterised by cortical myoclonic tremor usually from the second decade of life and overt myoclonic or generalised tonic-clonic seizures. Four independent loci have been implicated in FAME on chromosomes (chr) 2, 3, 5 and 8. Using whole genome sequencing and repeat primed PCR, we provide evidence that chr2-linked FAME (FAME2) is caused by an expansion of an ATTTC pentamer within the first intron of STARD7. The ATTTC expansions segregate in 158/158 individuals typically affected by FAME from 22 pedigrees including 16 previously reported families recruited worldwide. RNA sequencing from patient derived fibroblasts shows no accumulation of the AUUUU or AUUUC repeat sequences and STARD7 gene expression is not affected. These data, in combination with other genes bearing similar mutations that have been implicated in FAME, suggest ATTTC expansions may cause this disorder, irrespective of the genomic locus involvedSupplementary Information: Supplementary Data 1; Supplementary Data 2; Reporting Summary.NHMRC; Women’s and Children’s Hospital Research Foundation; Muir Maxwell Trust; Epilepsy Society; The European Fund for Regional Development; The province of Friesland, Dystonia Medical Research Foundation; Stichting Wetenschapsfonds Dystonie Vereniging; Fonds Psychische Gezondheid; Phelps Stichting; The Italian Ministry of Health; Istituto Superiore di Sanità, Italy; Undiagnosed Disease Network Italy; The Fondation maladies rares, University Hospital Essen and UK Department of Health’s NIHR.https://www.nature.com/ncommspm2020Neurolog

Phenotype associated with TAF2 biallelic mutations: a clinical description of four individuals and review of the literature

International audienceTranscription factor IID is a multimeric protein complex that is essential for the initiation of transcription by RNA polymerase II. One of its critical components, the TATA-binding protein-associated factor 2, is encoded by the gene TAF2. Pathogenic variants of this gene have been shown to be responsible for the Mental retardation, autosomal recessive 40 syndrome. This syndrome is characterized by severe intellectual disability, postnatal microcephaly, pyramidal signs and thin corpus callosum. Until now, only three families have been reported separately. Here we report four individuals, from two unrelated families, who present with severe intellectual disability and global developmental delay, postnatal microcephaly, feet deformities and thin corpus callosum and who carry homozygous TAF2 missense variants detected by Exome Sequencing. Taken together, our findings and those of previously reported subjects allow us to further delineate the clinical phenotype associated with TAF2 biallelic mutations

Combination of in silico and proteomic approaches to identify candidate genes responsible for the immunomodulatory properties of[i] Propionibacterium freudenreichii[/i]

Recent works have revealed highly strain-dependent immunomodulatory properties in Propionibacterium freudenreichii, a cheese starter bacterium. The underlying mechanisms are unknown but preliminary attempts have shown that surface components are involved in these immunomodulatory effects. The most promising strains/components exert promising anti-inflammatory effects through induction of regulatory cytokines.The aim of this work is to identify key surface components of Propionibacterium freudenreichii responsible for the immunomodulatory properties. 12 strains of Propionibacterium freudenreichii were sorted according to their high or low regulatory properties, measured by induction of IL-10 cytokine in peripheral blood mononuclear cells stimulated by equal amounts of the different P. freudenreichii strains. These strains were sequenced by Illumina paired-end sequencing and de novo assembled using Velvet software. Genome sequences were automatically and manually annotated on the INRA-AGMIAL platform. Correlations are studied both (i) between quantitative traits (high or low induction of IL-10) and genotypic properties, namely presence/lack of proteins predicted in silico and (ii) between quantitative traits (high or low induction of IL-10) and in vitro data (surface exposed proteins). In both cases, we account for the phylogenetic inertia induced by the shared evolutionary history of the strains. For in vitro data, three methods (shedding using guanidium chloride, surface labeling using CyDye cyanine and shaving using trypsine combined with mass spectrometry) are used to identify P. freudenreichii surface proteins. Statistical analysis is used to highlight which of these proteins can potentially be involved in immunomodulation, comparing the proteins identified and IL-10 induction. Statistical analyses of both in vitro and in silico approaches based on genome sequencing reveal targets potentially involved in immunomodulation. Both strategies are complementary. Proteomic approach followed by statistical analysis allow the identification of surface exposed proteins associated with immunomodulation, whereas in silico approach point out the mechanisms explaining the presence/ absence of the candidates on bacterial surface.Genome sequences, either directly used (in silico approach) or used as database (in vitro approach) determine key genes involved in the studied phenotypes. Candidate genes identified in the present study will be further characterized and their role in immunomodulation confirmed by knock out or overexpression of selected genes

Combination of in silico and proteomic approaches to identify candidate genes responsible for the immunomodulatory properties of Propionibacterium freudenreichii

Recent works have revealed highly strain-dependent immunomodulatory properties in Propionibacterium freudenreichii, a cheese starter bacterium (Foligné et al. 2010). The underlying mechanisms are unknown but preliminary attempts have shown that surface components are involved in these immunomodulatory effects. The most promising strains/components exert promising anti-inflammatory effects through induction of regulatory cytokines. The aim of this work is to identify key surface components of Propionibacterium freudenreichii responsible for the immunomodulatory properties. 12 strains of Propionibacterium freudenreichii were sorted according to their high or low regulatory properties, measured by induction of IL-10 cytokine in peripheral blood mononuclear cells stimulated by equal amounts of the different P. freudenreichii strains. These strains were sequenced by Illumina paired-end sequencing and de novo assembled using Velvet software. Genome sequences were automatically and manually annotated on the INRA-AGMIAL platform. Correlations are studied both (i) between quantitative traits (high or low induction of IL-10) and genotypic properties, namely presence/lack of in silico predicted proteins and (ii) between quantitative traits (high or low induction of IL-10) and in vitro data (surface exposed proteins). In both cases, we account for the phylogenetic inertia induced by the shared evolutionary history of the strains. For in vitro data, three methods (shedding using guanidium chloride, surface labeling using CyDye cyanine and shaving using trypsine combined with mass spectrometry) are used to identify P. freudenreichii surface proteins. Statistical analysis is used to highlight which of these proteins can potentially be involved in immunomodulation, comparing the proteins identified and IL-10 induction. Statistical analyses of both in vitro and in silico approaches based on genome sequencing reveal targets potentially involved in immunomodulation. Both strategies are complementary. Proteomic approach followed by statistical analysis allow the identification of surface exposed proteins associated with immunomodulation, whereas in silico approach point out the mechanisms explaining the presence/ absence of the candidates on bacterial surface. Genome sequences, either directly used (in silico approach) or used as database (in vitro approach) determine key genes involved in the studied phenotypes. Candidate genes identified in the present study will be further characterized and their role in immunomodulation confirmed by knock out or overexpression of selected genes

Combination of in silico and proteomic approaches to identify candidate genes responsible for the immunomodulatory properties of Propionibacterium freudenreichii

Recent works have revealed highly strain-dependent immunomodulatory properties in Propionibacterium freudenreichii, a cheese starter bacterium (Foligné et al. 2010). The underlying mechanisms are unknown but preliminary attempts have shown that surface components are involved in these immunomodulatory effects. The most promising strains/components exert promising anti-inflammatory effects through induction of regulatory cytokines. The aim of this work is to identify key surface components of Propionibacterium freudenreichii responsible for the immunomodulatory properties. 12 strains of Propionibacterium freudenreichii were sorted according to their high or low regulatory properties, measured by induction of IL-10 cytokine in peripheral blood mononuclear cells stimulated by equal amounts of the different P. freudenreichii strains. These strains were sequenced by Illumina paired-end sequencing and de novo assembled using Velvet software. Genome sequences were automatically and manually annotated on the INRA-AGMIAL platform. Correlations are studied both (i) between quantitative traits (high or low induction of IL-10) and genotypic properties, namely presence/lack of in silico predicted proteins and (ii) between quantitative traits (high or low induction of IL-10) and in vitro data (surface exposed proteins). In both cases, we account for the phylogenetic inertia induced by the shared evolutionary history of the strains. For in vitro data, three methods (shedding using guanidium chloride, surface labeling using CyDye cyanine and shaving using trypsine combined with mass spectrometry) are used to identify P. freudenreichii surface proteins. Statistical analysis is used to highlight which of these proteins can potentially be involved in immunomodulation, comparing the proteins identified and IL-10 induction. Statistical analyses of both in vitro and in silico approaches based on genome sequencing reveal targets potentially involved in immunomodulation. Both strategies are complementary. Proteomic approach followed by statistical analysis allow the identification of surface exposed proteins associated with immunomodulation, whereas in silico approach point out the mechanisms explaining the presence/ absence of the candidates on bacterial surface. Genome sequences, either directly used (in silico approach) or used as database (in vitro approach) determine key genes involved in the studied phenotypes. Candidate genes identified in the present study will be further characterized and their role in immunomodulation confirmed by knock out or overexpression of selected genes