Unknown,Transcriptomics,Genomics,Proteomics

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Bodo saltans RNAseq

ABSTRACT: Abstract: The Kinetoplastida (Euglenozoa) are unicellular flagellates that include the trypanosomatid parasites, most notably Trypanosoma brucei, T.cruzi and Leishmania spp. These organisms cause substantial mortality and morbidity in humans and their livestock worldwide as the causative agents of African sleeping sickness, Chagas disease and leishmaniasis respectively. Draft genome sequences are available for several species of both Trypanosoma and Leishmania. Bodo saltans is a free-living heterotroph found worldwide in freshwater and marine habitats, and it is among the closest bodonid relatives of the trypanosomatids. The purpose of a B. saltans genome sequence is to provide an 'out-group' for comparative genomic analysis of the trypanosomatid parasites. It will provide a model of the ancestral trypanosomatid to distinguish those derived parts of the parasite genomes (i.e., unique trypanosomatid adaptations) from those which are a legacy of the free-living ancestor. To aid annotation of the B.saltans genome sequence, total genomic RNA was extracted on four occasions from the total cellular mass of 160ml of B.saltans cell culture, for the purposes of transcription profiling by high throughput sequencing. Cells were unmodified. B.saltans cells were grown in water at 4oC. Total genomic RNA was extracted from a cell pellet using TRIZOL reagent and ethanol precipitated. Poly A+ mRNA was purified from total RNA using oligo dT dyna bead selection and libraries were created using the Illumina RNA-seq protocol. The samples were sequenced on an Illumina HiSeq 2000. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/

INSTRUMENT(S): Illumina HiSeq 2000

ORGANISM(S): Bodo saltans

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PROVIDER: E-ERAD-88 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Protein-coding genes in kinetoplastid protists are transcribed from polycistronic arrays, yielding RNA precursors which are processed to form mature transcripts bearing a 5M-bM-^@M-^Y spliced leader (SL) and 3M-bM-^@M-^Y poly(A) tract. Regions of transcription initiation and termination lack known eukaryotic promoter and terminator elements, and current data suggest that transcription is instead regulated predominantly through epigenetic mechanisms. Several epigenetic marks, including histone modifications, histone variants, and an atypical DNA modification known as base J have been localized to regions of transcription initiation or termination in Trypanosoma brucei, Trypanosoma cruzi, and/or Leishmania major. Despite this conservation, the phenotypes of base J mutants vary significantly across trypanosomatids, suggesting that the specific epigenetic networks governing transcription initiation and termination have diverged significantly during evolution. In this light, we sought to characterize and compare the roles of the histone variants H2A.Z, H2B.V, and H3.V in L. major. As in T. brucei, the histone variants H2A.Z and H2B.V were shown to be essential in L. major using a powerful quantitative plasmid segregation-based test. In contrast and again similar to T. brucei, H3.V is not essential in Leishmania as H3.V-null lines grew normally, resembled WT, and remained infectious. Using SL-primed RNA-seq, we found that H3.V-null parasites have steady-state transcript levels comparable to WT parasites and display no defects in the efficiency of transcription termination at convergent strand switch regions (SSRs). Our results show a conservation of histone variant phenotypes between L. major and T. brucei, in contrast to the phenotypes associated with the epigenetic DNA base J modification. Total RNA from Four LmjF samples were analyzed using RNA-Seq. One of them is wildtype parasites, one is single knockout for H3V gene and two independent double knockouts for H3V gene.

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Bodo saltans RNAseq

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