Project description:A maturation, sex and lunar phase locked Platynereis head proteome was generated by sampling six heads per biological replicate from male and female Platynereis at defined stages of sexual maturation (immature, premature and mature) at two different moon phases; New moon and Free-running Full moon, meaning the animals would expect a nocturnal light stimulus, Full moon, which was not given. From these heads RNA and protein were extracted simultaneously and used for both transcriptomics (ENA accession no. PRJEB27496) and quantitative proteomics. In total 2290 proteins were identified with two unique peptides as requirement; of these, 1064 proteins were found to be quantifiable (identification in two out of three biological replicates and three out of five technical replicates). Relative quantification revealed that the majority of quantifiable proteins changed during maturation (693 out of 1064 proteins), sexual differences accounted for only 17 differentially expressed proteins, whereas lunar phase differences showed 261 differentially expressed proteins. This finding is in contrast to what was found on the transcriptome levels where differences in between the lunar phases only accounted for only 64 out of 52059 transcripts. To confirm this high degree of lunar regulated proteins a second proteome from immature animals at New moon and Free-running Full moon was undertaken. In this second set, we identified 2095 proteins of which 1671 were considered quantifiable. Of these 173 proteins showed significant differences between the New moon and the Free running full moon phase, thus confirming the previous results of a higher regulation on the protein compared to the transcript levels.
2019-02-19 | PXD010532 | Pride
Project description:NRPS adenylation domain amplicon data from wheat rhizosphere in drought experiments
Project description:Early development depends heavily on accurate control of maternally inherited mRNAs, and yet it remains unknown how maternal microRNAs (miRNAs) are regulated during maternal to zygotic transition (MZT). We here find that maternal miRNAs are highly adenylated at their 3' ends in mature oocytes and early embryos. Pervasive adenylation is observed in oocytes of fly, sea urchin and mouse, indicating that maternal miRNA adenylation may be widely conserved in animals. We identify Wispy as the enzyme responsible for miRNA adenylation in flies. Wispy is known to be expressed specifically in oocytes and early embryos and function as a noncanonical poly(A) polymerase. Knockout of wispy abrogates miRNA adenylation and induces miRNA accumulation in fly eggs whereas overexpression of Wispy increases adenylation and reduces miRNA levels in S2 cells. Adenylation occurs on both the 5p and 3p miRNAs, indicating that Wispy acts on miRNAs after Dicer processing. We further find that Wispy interacts with Ago1 through protein-protein interaction, which may allow the effective and selective adenylation of miRNAs. Thus, adenylation may contribute to the clearance of maternally deposited miRNAs during MZT. Our work provides the first mechanistic insights into the regulation of maternal miRNAs and illustrates the importance of RNA tailing in development.
Project description:The Poly(A)-Tail focused RNA-seq, or PAT-seq approach, is an affordable and efficient tool for the measure of 3’UTR dynamics. We show here that PAT-seq returns (i) digital gene expression, (ii) polyadenylation site usage within and between samples, including alternative adenylation, and (iii) the polyadenylation-state the transcriptome. PAT-seq differs from previous 3’ focused RNA-seq methods in that it strictly depends on native 3’ adenylation within total RNA samples and thus removes the need for ribosome depletion and, that the full native poly(A)-tail is included in the sequencing libraries. Limited RNase digestion combined with size selection and directional sequencing mean that deep-sequencing reads map to within ~50-100 bases of adenylation sites and run from unique sequence into adenosine homopolymers. Here, total RNA samples from budding yeast cells were analyzed to highlight the changes in gene expression and adenylation-state of the transcriptome in response to loss of the deadenylase Ccr4. Furthermore, concordant changes to gene expression and adenylation-state were demonstrated in a classic Crabtree-Warburg metabolic shift. Because all adenylated RNA are interrogated by the PAT-seq approach, alternative adenylation sites, long noncoding RNA and other non-coding RNA and RNA decay intermediates were also identified.
Project description:Early development depends heavily on accurate control of maternally inherited mRNAs, and yet it remains unknown how maternal microRNAs (miRNAs) are regulated during maternal to zygotic transition (MZT). We here find that maternal miRNAs are highly adenylated at their 3' ends in mature oocytes and early embryos. Pervasive adenylation is observed in oocytes of fly, sea urchin and mouse, indicating that maternal miRNA adenylation may be widely conserved in animals. We identify Wispy as the enzyme responsible for miRNA adenylation in flies. Wispy is known to be expressed specifically in oocytes and early embryos and function as a noncanonical poly(A) polymerase. Knockout of wispy abrogates miRNA adenylation and induces miRNA accumulation in fly eggs whereas overexpression of Wispy increases adenylation and reduces miRNA levels in S2 cells. Adenylation occurs on both the 5p and 3p miRNAs, indicating that Wispy acts on miRNAs after Dicer processing. We further find that Wispy interacts with Ago1 through protein-protein interaction, which may allow the effective and selective adenylation of miRNAs. Thus, adenylation may contribute to the clearance of maternally deposited miRNAs during MZT. Our work provides the first mechanistic insights into the regulation of maternal miRNAs and illustrates the importance of RNA tailing in development. MiRNA expression and modification profile during early embryo development of fruit fly and zebra fish using high throughput sequencing
Project description:This study is part of MD Anderson Cancer Center CRC Moon Shot. We used single cell RNAsequencing (scRNA-seq) to analyze the diversity of CRC.
Project description:ChIP-seq analysis of SNAIL binding sites in RH30 cells was performed to discover novel SNAIL binding sites in rhabdmyosarcoma cells.
Project description:In skeletal myogenesis, the transcription factor MyoD activates distinct transcriptional programs in progenitors compared to terminally differentiated cells. Using ChIP-seq and gene expression analyses, we show that in primary myoblasts, Snail-HDAC1/2 repressive complex bind and exclude MyoD from its targets. Notably, Snail binds E-box motifs that are G/C-rich in their central dinucleotides, and such sites are almost exclusively associated with genes expressed during differentiation. By contrast, Snail does not bind the A/T-rich E-boxes associated with MyoD targets in myoblasts. Thus, Snai1-HDAC1/2 prevents MyoD occupancy on differentiation-specific regulatory elements and the change from Snail- to MyoD-binding often results in enhancer switching during differentiation. Furthermore, we show that a regulatory network involving Myogenic Regulatory Factors (MRFs), Snail/2, miR-30a and miR-206 acts as a molecular switch that controls entry into myogenic differentiation. Together, these results reveal a regulatory paradigm that directs distinct gene expression programs in progenitors versus terminally differentiated cells. Genome wide binding sites of various transcription factors and chromatin modifiers in muscle cells