Project description:Adaptation of eukaryotic cells to anaerobic condition is reflected by deep changes in mitochondrial metabolism and their functional reduction. The most modified types of mitochondria are hydrogenosomes that generate molecular hydrogen with concomitant ATP synthesis and mitosome that completely lost energy metabolism. The reduction of mitochondria is associated with loss of peroxisomes that evolved from ER to compartmentalize pathways generating reactive oxygen species (ROS) and thus prevent cellular oxidative damage. Biogenesis and function of peroxisomes is tightly coupled with mitochondria. They share the fission machinery, pathways of oxidative metabolism, ROS scavenging, and metabolic products. The loss of peroxisomes in anaerobic eukaryotes with reduced mitochondria is thus not unexpected. Surprisingly, we identified peroxisomes in anaerobic, hydrogenosome bearing protist Mastigamoeba balamuthi. Initially, we identified conserved set of peroxisomal proteins peroxins that are required for protein import, peroxisomal growth and division. Key membrane associated peroxins (MbPex3, MbPex11, and MbPex14) were visualized in numerous vesicles that were distinct from hydrogenosomes, ER and Golgi body. Proteomic analysis of cellular fractions and prediction of peroxisomal targeting signals (PTS1/PTS2) allows identification of 51 putative peroxisomal matrix proteins. Expression of selected proteins in Saccharomyces cerevisiae revealed that they are specifically targeted to yeast peroxisomes. Matrix protein includes components of acyl CoA and carbohydrate metabolism, pyrimidine and CoA biosynthesis, whereas neither components of β-oxidation nor catalase were present. In conclusion, we identified new subclass of peroxisomes named “anaerobic” peroxisome that shifts the current paradigm and rises attention to reductive evolution of peroxisomes in anaerobic organisms.
Project description:E. histolytica is believed to be devoid of peroxisomes, like most anaerobic protists. In this work, we provided the first evidence that peroxisomes are present in E. histolytica, although the set of proteins (peroxins) responsible for peroxisome biogenesis was reduced to only seven members (Pex1, Pex6, Pex5, Pex11, Pex14, Pex16, and Pex19). Targeting matrix proteins to peroxisomes is reduced to the PTS1-dependent pathway mediated via the soluble Pex5 receptor, while the PTS2 receptor Pex7 is absent. Immunofluorescence microscopy showed that peroxisomal markers (Pex5, Pex14, Pex16, Pex19) are present in vesicles distinct from mitosomes, the endoplasmic reticulum and the endosome/phagosome system, except Pex11, which has dual localization in peroxisomes and mitosomes. Immunoelectron microscopy revealed that Pex14 localized to vesicles of approximately 90-100 nm in diameter. Proteomic analyses of affinity-purified peroxisomes and in silico PTS1 predictions provided datasets of 655 and 56 peroxisomal candidates, respectively; however, only six proteins were shared by both datasets, including myo-inositol dehydrogenase (myo-IDH). Peroxisomal NAD-dependent myo-IDH appeared to be a dimeric enzyme with high affinity to myo-inositol (Km 0,044 mM) and can utilize also scyllo-inositol, D-glucose and D-xylose as substrates. Phylogenetic analyses revealed that orthologs of myo-IDH with PTS1 are present in E. dispar, E. nutalli and E. moshkovskii but not in E. invadens, and form a monophyletic clade of mostly peroxisomal orthologs with free-living M. balamuthi and P. schiedti. The presence of peroxisomes in E. histolytica and other archamoebae breaks the paradigm of peroxisome absence in anaerobes and provides a new potential target for the development of antiparasitic drugs.
Project description:The DNA isolated from 44 either frozen or FFPE Neuroendocrine Neoplasm (NEN) was analysed by NGS, to identify genes more likely to be subject to sequence variations among 523 cancer-related ones.
Project description:Plasma DNA from 558 malignancies, 263 benign and borderline tumors and 367 healthy control samples were collected and subjected to random short-gun whole genome sequencing.
Project description:This study aims to investigate the DNA methylation patterns at transcription factor binding regions and their evolutionary conservation with respect to binding activity divergence. We combined newly generated bisulfite-sequencing experiments in livers of five mammals (human, macaque, mouse, rat and dog) and matched publicly available ChIP-sequencing data for five transcription factors (CEBPA, HNF4a, CTCF, ONECUT1 and FOXA1). To study the chromatin contexts of TF binding subjected to distinct evolutionary pressures, we integrated publicly available active promoter, active enhancer and primed enhancer calls determined by profiling genome wide patterns of H3K27ac, H3K4me3 and H3K4me1.