Project description:Interventions: ntestinal polyp gruop and colorectal cancer gruop:Nil
Primary outcome(s): bacteria;fungi;archaea;virus
Study Design: Factorial
Project description:Organisms of the third domain of life, the Archaea, share molecular characteristics both with bacteria and eukarya. These organisms attract scientific attention as research models for regulation and evolution of processes such as transcription, translation and RNA processing. We have reconstructed the primary transcriptome of Sulfolobus solfataricus P2, one of the most widely studied model archaeal organisms. Analysis of 625 million bases of sequenced cDNAs yielded a single-bp resolution map of transcription start sites and operon structures for more than 1000 transcriptional units. The analysis led to the discovery of 310 expressed non-coding RNAs, with an extensive expression of overlapping cis-antisense transcripts to a level unprecedented in any bacteria or archaea but resembling that of eukaryotes. As opposed to bacterial transcripts, most Sulfolobus transcripts completely lack 5' UTR sequences, suggesting that mRNA/ncRNA interactions differ between bacteria and archaea. The data also reveal internal hotspots for transcript cleavage linked to RNA degradation, and predict sequence motifs that promote RNA destabilization. This study emphasizes the importance of transcriptome sequencing as a key tool for understanding the mechanisms and extent of RNA-based regulation for bacteria and archaea. 5 samples of cDNA sequencing (2 of these are replicates), and 3 samples of RACE-cDNA sequencing (described in the samples section).
Project description:Organisms of the third domain of life, the Archaea, share molecular characteristics both with bacteria and eukarya. These organisms attract scientific attention as research models for regulation and evolution of processes such as transcription, translation and RNA processing. We have reconstructed the primary transcriptome of Sulfolobus solfataricus P2, one of the most widely studied model archaeal organisms. Analysis of 625 million bases of sequenced cDNAs yielded a single-bp resolution map of transcription start sites and operon structures for more than 1000 transcriptional units. The analysis led to the discovery of 310 expressed non-coding RNAs, with an extensive expression of overlapping cis-antisense transcripts to a level unprecedented in any bacteria or archaea but resembling that of eukaryotes. As opposed to bacterial transcripts, most Sulfolobus transcripts completely lack 5' UTR sequences, suggesting that mRNA/ncRNA interactions differ between bacteria and archaea. The data also reveal internal hotspots for transcript cleavage linked to RNA degradation, and predict sequence motifs that promote RNA destabilization. This study emphasizes the importance of transcriptome sequencing as a key tool for understanding the mechanisms and extent of RNA-based regulation for bacteria and archaea.
Project description:This study evaluated the ammonium oxidizing communities (COA) associated with a potato crop (Solanum phureja) rhizosphere soil in the savannah of Bogotá (Colombia) by examining the presence and abundance of amoA enzyme genes and transcripts by qPCR and next-generation sequence analysis. amoA gene abundance could not be quantified by qPCR due to problems inherent in the primers; however, the melting curve analysis detected increased fluorescence for Bacterial communities but not for Archaeal communities. Transcriptome analysis by next-generation sequencing revealed that the majority of reads mapped to ammonium-oxidizing Archaea, suggesting that this activity is primarily governed by the microbial group of the Crenarchaeota phylum. In contrast,a lower number of reads mapped to ammonia-oxidizing bacteria.
Project description:Structure probing combined with next-generation sequencing (NGS) has provided novel insights into RNA structure-function relationships. To date such studies have focused largely on bacteria and eukaryotes, with little attention given to the third domain of life, archaea. Furthermore, functional RNAs have not been extensively studied in archaea, leaving open questions about RNA structure and function within this domain of life. With archaeal species being diverse and having many similarities to both bacteria and eukaryotes, the archaea domain has the potential to be an evolutionary bridge. In this study, we introduce a method for probing RNA structure in vivo in the archaea domain of life. We investigated the structure of ribosomal RNA (rRNA) from Methanosarcina acetivorans, a well-studied anaerobic archaeal species, grown with either methanol or acetate. After probing the RNA in vivo with dimethyl sulfate (DMS), Structure-seq2 libraries were generated, sequenced, and analyzed. We mapped the reactivity of DMS onto the secondary structure of the ribosome, which we determined independently with comparative analysis, and confirmed the accuracy of DMS probing in M. acetivorans. Accessibility of the rRNA to DMS in the two carbon sources was found to be quite similar, although some differences were found. Overall, this study establishes the Structure-seq2 pipeline in the archaea domain of life and informs about ribosomal structure within M. acetivorans.
Project description:Histone proteins have traditionally been thought to be restricted to eukaryotes and most archaea, with eukaryotic nucleosomal histones deriving from their archaeal ancestors. In contrast, bacteria lack histones as a rule. However, in recent years histone proteins have been identified in a few bacterial clades, in particular the phylum Bdellovibrionota, and these histones have been proposed to exhibit a range of divergent features compared to histones in archaea and eukaryotes. However, no experimental functional genomic studies of the properties of Bdellovibrionota chromatin have been carried out. In this work, we map the landscape of chromatin accessibility, active transcription and three-dimensional genome organization in a member of Bdellovibrionota (a Bacteriovorax strain). We find that Bacteriovorax chromatin is characterized by preferential accessibility around promoter regions, similar to what is observed in eukaryotes with compact genomes such as yeast, and also to some archaea. As in eukaryotes, chromatin accessibility positively correlates with gene expression. Mapping active transcription through single-strand DNA (ssDNA) profiling revealed that Bacteriovorax promoters exhibit very strong polymerase pausing, unlike in yeast, but similar to the state of mammalian and fly promoters. Finally, the Bacteriovorax genome exists in a three-dimensional (3D) conformation analogous to that of other bacteria without histones, organized by the parABS system and along the axis defined by replication origin and termination regions. These results provide a foundation for understanding the chromatin biology of the unique Bdellovibrionota bacteria and the deep evolution of chromatin organization across the tree of life.