Project description:We provide sequences of maize chlroplast RNAs associated with non-PPR editing factors through immunoprecipitation. The accumulated sequences indicate expansive role in RNA processing and potential associations of OZ1-ORRM1 complexes with the ribosome and less efficiently translated mRNAs.
Project description:ngs2019_18_eplus-eplus-search for mitochondrial editing defect in an arabidopsis PPR mutant Annotation, RNA/Small-RNA quantification: editing quantification. The Mito samples were first enriched with mitochondria by a series of multi-speed centrifugations after grinding with mortar at 4°C.
Project description:ngs2018_04_half_edit-half_edit - Is there some transcriptomic defects in these different PPR KO mutants? - Identification of RNA editing defects in 3 differents KO mutants for E+ PPR. Results will be compared to different predictive methods in order to find out which one is the more accurate. Also looking for other transcriptomic defects in a pure-PPR.
Project description:Pentatricopeptide repeat (PPR) proteins, which are characterized by tandem 30-40 amino acid sequence motifs, constitute a large gene family in plants. These known PPR proteins have been identified to play important roles in organellar RNA metabolism and plant development in Arabidopsis and rice. However, functions of PPR genes in woody species remain still largely unknown. Here, we identified and characterized a total of 626 PPR genes containing PPR motifs in the poplar genome. A comprehensive genome-wide analysis of the poplar PPR gene family was performed, including chromosomal location, phylogenetic relationships, gene duplication. Transcriptomic analyses identified that 154 of the PtrPPR genes were induced by biotic and abiotic treatments, including Marssonina brunnea, salicylic acid (SA), methyl jasmonate (MeJA), wounding, cold and salinity. Quantitative RT-PCR analysis further confirmed the expression profiles of 11 PtrPPR genes under different stresses. Our results contribute to a more comprehensive understanding the roles of PPR proteins and provided an insight for improving the stress tolerance in poplar.
Project description:RNA editing is converting hundreds of cytosines into uridines during organelle gene expression of land plants. The pentatricopeptide repeat (PPR) proteins are at the core of this posttranscriptional RNA modification. Even if a PPR protein defines the editing site, a DYW domain of the same or another PPR protein is believed to catalyze the deamination. To give insight into the organelle RNA editosome, we performed tandem affinity purification of the plastidial CHLOROPLAST BIOGENESIS 19 (CLB19) PPR editing factor. Two PPR proteins, dually targeted to mitochondria and chloroplasts, were identified as potential partners of CLB19. These two proteins, a P-type PPR and a member of a small PPR-DYW subfamily, were shown to interact in yeast. Insertional mutations resulted in embryo lethality that could be rescued by embryo-specific complementation. A transcriptome analysis of these complemented plants showed major editing defects in both organelles with a very high PPR type specificity, indicating that the two proteins are core members of E+-type PPR editosomes.
Project description:To demonstrate RIPSeeker program that is developed for RIP-seq analyses, we generated RIP-seq data corresponding to the protein CCNT1 in HEK293 cell line using standard RIP-seq protocols described in Zhao et al., (2010). We performed two in-house RIP-seq experiments both for CCNT1 in human HEK293 cells. Briefly, we generated tagged CCNT1 using a triple tag system that supports lentiviral stable expression and mammalian affinity purification (MAPLE) Mak et al (2010). The HEK293 cells stably expressing tagged CCNT1 was purified by M2 agarose beads, followed by RNA extraction by Trizol. The library synthesis was carried out according to the RIP-seq protocol described in Zhao et al., (2010) except that one of the two experiments was done with non-strand-specific sequencing.
Project description:Sox2 is a master transcriptional regulator of embryonic development. Having found that Sox2 interacts with RNA-binding proteins, we designed an experiment to discover RNAs associated with Sox2 and other pluripotency factors. Briefly, we used RNA immunoprecipitation followed by high-throughput sequencing (RIP-seq) to sequence transcriptomes enriched for RNAs associated with Klf4, Nanog, Oct4, Sox2 and Suz12. For completeness, this submission includes all RIP-seq data from this study, although some of the data was only used for exploratory analyses. Such analysis indicated that it was important to include input samples for each of the the cell lines that were to be compared, to account for differences in gene expression (e.g. between J1-birA with and without bioSox2), and that overnight IP was more informative than 3 h IP. Therefore, the analysis described in the associated manuscript used samples from experiment batch 3 and 4 only (samples 11-18).
Project description:In a recent study, we showed that a T-DNA insertional mutation in a mitochondrial PPR protein, POCO1, led to the earlier floral transition (Emami and Kempken 2019). We used RNA-seq analysis to provide an overview of the global transcriptome changes in poco1 mutant during different developmental stages.