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Systematic imaging reveals features and changing localization of mRNAs in Drosophila development.

Jambor H, Surendranath V, Kalinka AT, Mejstrik P, Saalfeld S, Tomancak P - Elife (2015)

Bottom Line: We combined transcriptomics and systematic imaging to determine the tissue-specific expression and subcellular distribution of 5862 mRNAs during Drosophila oogenesis. mRNA localization is widespread in the ovary and detectable in all of its cell types-the somatic epithelial, the nurse cells, and the oocyte.Genes defined by a common RNA localization share distinct gene features and differ in expression level, 3'UTR length and sequence conservation from unlocalized mRNAs.Comparison of mRNA localizations in different contexts revealed that localization of individual mRNAs changes over time in the oocyte and between ovarian and embryonic cell types.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.

ABSTRACT
mRNA localization is critical for eukaryotic cells and affects numerous transcripts, yet how cells regulate distribution of many mRNAs to their subcellular destinations is still unknown. We combined transcriptomics and systematic imaging to determine the tissue-specific expression and subcellular distribution of 5862 mRNAs during Drosophila oogenesis. mRNA localization is widespread in the ovary and detectable in all of its cell types-the somatic epithelial, the nurse cells, and the oocyte. Genes defined by a common RNA localization share distinct gene features and differ in expression level, 3'UTR length and sequence conservation from unlocalized mRNAs. Comparison of mRNA localizations in different contexts revealed that localization of individual mRNAs changes over time in the oocyte and between ovarian and embryonic cell types. This genome scale image-based resource (Dresden Ovary Table, DOT, http://tomancak-srv1.mpi-cbg.de/DOT/main.html) enables the transition from mechanistic dissection of singular mRNA localization events towards global understanding of how mRNAs transcribed in the nucleus distribute in cells.

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The transcriptome shows little variation over the course of oogenesis.(A) Scatterplot showing high correlation (Pearson Correlation 0.71) between RNAseq and 3Pseq sequencing results. (B) Stage-specific sequencing reveals that ∼5500 genes (grey) were detected by both mRNA sequencing (RNAseq) and 3′prime end sequencing (3Pseq). Additional 1000–2000 mRNAs were captures with either RNAseq (green) or 3Pseq (red) only, suggesting that at each time point about half of the D. melanogaster genome was expressed. (C) Venn diagram showing that most genes, 85%, are continuously expressed as they are detectable at each time point of oogenesis (3Pseq: red; RNAseq: green). (D) Across oogenesis RNA levels were highly correlated, suggesting minimal changes in the stage-specific transcriptomes (Pair-wise correlation, Pearson Correlation: 0.77; significantly up/down regulated genes shown in black: p-value adjusted for multiple testing <0.1, see also Figure 5B). (E) GO-term analysis of genes significantly up- (arrow up) or down- (arrow down) regulated during oogenesis/early embryogenesis. Particularly genes encoding components of the extra-embryonic layers (vitelline membrane, ECM, cuticle) changed expression levels. (F) nanos mRNA significantly changes gene expression from early to full ovaries measured by RNAseq (green) and 3Pseq (red). Below: gene model. (G) Boxplot showing that the vast majority of genes expressed only one 3′UTR form during oogenesis, suggesting low prevalence of alternative polyadenylation. (H) Correlation of expressed transcripts (measured by RNAseq) revealed that only few genes significantly (shown in black) changed the expressed isoform during oogenesis (see also Figure 5C). (I) Only few subcellular localized mRNAs showed significant changes in transcript expression (see H). (J) The number of transcripts expressed per gene did not change during oogenesis did not differ between ubiquitous and subcellular gene sets (highlighted: anterior [red] and posterior [blue] genes).DOI:http://dx.doi.org/10.7554/eLife.05003.016
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fig5s1: The transcriptome shows little variation over the course of oogenesis.(A) Scatterplot showing high correlation (Pearson Correlation 0.71) between RNAseq and 3Pseq sequencing results. (B) Stage-specific sequencing reveals that ∼5500 genes (grey) were detected by both mRNA sequencing (RNAseq) and 3′prime end sequencing (3Pseq). Additional 1000–2000 mRNAs were captures with either RNAseq (green) or 3Pseq (red) only, suggesting that at each time point about half of the D. melanogaster genome was expressed. (C) Venn diagram showing that most genes, 85%, are continuously expressed as they are detectable at each time point of oogenesis (3Pseq: red; RNAseq: green). (D) Across oogenesis RNA levels were highly correlated, suggesting minimal changes in the stage-specific transcriptomes (Pair-wise correlation, Pearson Correlation: 0.77; significantly up/down regulated genes shown in black: p-value adjusted for multiple testing <0.1, see also Figure 5B). (E) GO-term analysis of genes significantly up- (arrow up) or down- (arrow down) regulated during oogenesis/early embryogenesis. Particularly genes encoding components of the extra-embryonic layers (vitelline membrane, ECM, cuticle) changed expression levels. (F) nanos mRNA significantly changes gene expression from early to full ovaries measured by RNAseq (green) and 3Pseq (red). Below: gene model. (G) Boxplot showing that the vast majority of genes expressed only one 3′UTR form during oogenesis, suggesting low prevalence of alternative polyadenylation. (H) Correlation of expressed transcripts (measured by RNAseq) revealed that only few genes significantly (shown in black) changed the expressed isoform during oogenesis (see also Figure 5C). (I) Only few subcellular localized mRNAs showed significant changes in transcript expression (see H). (J) The number of transcripts expressed per gene did not change during oogenesis did not differ between ubiquitous and subcellular gene sets (highlighted: anterior [red] and posterior [blue] genes).DOI:http://dx.doi.org/10.7554/eLife.05003.016

Mentions: Based on our in situ hybridization screen, we identified 3475 mRNAs as being expressed and most of these mRNAs were also detectable by RNA sequencing. Both sequencing techniques were in good agreement with each other (Figure 1A, Figure 2—figure supplement 1A, Figure 5—figure supplement 1A). Of the expressed genes, 64% showed ubiquitous mRNA distribution in ovary cells throughout oogenesis (ubiquitous), but we also observed mRNA expressions restricted to subsets of cells (cellular) and mRNAs that asymmetrically localized in the cytoplasm (subcellular) or to the nuclei of cells (nuclear).10.7554/eLife.05003.003Figure 1.Summary of the fluorescent in situ hybridization (FISH) screen in ovaries.


Systematic imaging reveals features and changing localization of mRNAs in Drosophila development.

Jambor H, Surendranath V, Kalinka AT, Mejstrik P, Saalfeld S, Tomancak P - Elife (2015)

The transcriptome shows little variation over the course of oogenesis.(A) Scatterplot showing high correlation (Pearson Correlation 0.71) between RNAseq and 3Pseq sequencing results. (B) Stage-specific sequencing reveals that ∼5500 genes (grey) were detected by both mRNA sequencing (RNAseq) and 3′prime end sequencing (3Pseq). Additional 1000–2000 mRNAs were captures with either RNAseq (green) or 3Pseq (red) only, suggesting that at each time point about half of the D. melanogaster genome was expressed. (C) Venn diagram showing that most genes, 85%, are continuously expressed as they are detectable at each time point of oogenesis (3Pseq: red; RNAseq: green). (D) Across oogenesis RNA levels were highly correlated, suggesting minimal changes in the stage-specific transcriptomes (Pair-wise correlation, Pearson Correlation: 0.77; significantly up/down regulated genes shown in black: p-value adjusted for multiple testing <0.1, see also Figure 5B). (E) GO-term analysis of genes significantly up- (arrow up) or down- (arrow down) regulated during oogenesis/early embryogenesis. Particularly genes encoding components of the extra-embryonic layers (vitelline membrane, ECM, cuticle) changed expression levels. (F) nanos mRNA significantly changes gene expression from early to full ovaries measured by RNAseq (green) and 3Pseq (red). Below: gene model. (G) Boxplot showing that the vast majority of genes expressed only one 3′UTR form during oogenesis, suggesting low prevalence of alternative polyadenylation. (H) Correlation of expressed transcripts (measured by RNAseq) revealed that only few genes significantly (shown in black) changed the expressed isoform during oogenesis (see also Figure 5C). (I) Only few subcellular localized mRNAs showed significant changes in transcript expression (see H). (J) The number of transcripts expressed per gene did not change during oogenesis did not differ between ubiquitous and subcellular gene sets (highlighted: anterior [red] and posterior [blue] genes).DOI:http://dx.doi.org/10.7554/eLife.05003.016
© Copyright Policy
Related In: Results  -  Collection

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fig5s1: The transcriptome shows little variation over the course of oogenesis.(A) Scatterplot showing high correlation (Pearson Correlation 0.71) between RNAseq and 3Pseq sequencing results. (B) Stage-specific sequencing reveals that ∼5500 genes (grey) were detected by both mRNA sequencing (RNAseq) and 3′prime end sequencing (3Pseq). Additional 1000–2000 mRNAs were captures with either RNAseq (green) or 3Pseq (red) only, suggesting that at each time point about half of the D. melanogaster genome was expressed. (C) Venn diagram showing that most genes, 85%, are continuously expressed as they are detectable at each time point of oogenesis (3Pseq: red; RNAseq: green). (D) Across oogenesis RNA levels were highly correlated, suggesting minimal changes in the stage-specific transcriptomes (Pair-wise correlation, Pearson Correlation: 0.77; significantly up/down regulated genes shown in black: p-value adjusted for multiple testing <0.1, see also Figure 5B). (E) GO-term analysis of genes significantly up- (arrow up) or down- (arrow down) regulated during oogenesis/early embryogenesis. Particularly genes encoding components of the extra-embryonic layers (vitelline membrane, ECM, cuticle) changed expression levels. (F) nanos mRNA significantly changes gene expression from early to full ovaries measured by RNAseq (green) and 3Pseq (red). Below: gene model. (G) Boxplot showing that the vast majority of genes expressed only one 3′UTR form during oogenesis, suggesting low prevalence of alternative polyadenylation. (H) Correlation of expressed transcripts (measured by RNAseq) revealed that only few genes significantly (shown in black) changed the expressed isoform during oogenesis (see also Figure 5C). (I) Only few subcellular localized mRNAs showed significant changes in transcript expression (see H). (J) The number of transcripts expressed per gene did not change during oogenesis did not differ between ubiquitous and subcellular gene sets (highlighted: anterior [red] and posterior [blue] genes).DOI:http://dx.doi.org/10.7554/eLife.05003.016
Mentions: Based on our in situ hybridization screen, we identified 3475 mRNAs as being expressed and most of these mRNAs were also detectable by RNA sequencing. Both sequencing techniques were in good agreement with each other (Figure 1A, Figure 2—figure supplement 1A, Figure 5—figure supplement 1A). Of the expressed genes, 64% showed ubiquitous mRNA distribution in ovary cells throughout oogenesis (ubiquitous), but we also observed mRNA expressions restricted to subsets of cells (cellular) and mRNAs that asymmetrically localized in the cytoplasm (subcellular) or to the nuclei of cells (nuclear).10.7554/eLife.05003.003Figure 1.Summary of the fluorescent in situ hybridization (FISH) screen in ovaries.

Bottom Line: We combined transcriptomics and systematic imaging to determine the tissue-specific expression and subcellular distribution of 5862 mRNAs during Drosophila oogenesis. mRNA localization is widespread in the ovary and detectable in all of its cell types-the somatic epithelial, the nurse cells, and the oocyte.Genes defined by a common RNA localization share distinct gene features and differ in expression level, 3'UTR length and sequence conservation from unlocalized mRNAs.Comparison of mRNA localizations in different contexts revealed that localization of individual mRNAs changes over time in the oocyte and between ovarian and embryonic cell types.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.

ABSTRACT
mRNA localization is critical for eukaryotic cells and affects numerous transcripts, yet how cells regulate distribution of many mRNAs to their subcellular destinations is still unknown. We combined transcriptomics and systematic imaging to determine the tissue-specific expression and subcellular distribution of 5862 mRNAs during Drosophila oogenesis. mRNA localization is widespread in the ovary and detectable in all of its cell types-the somatic epithelial, the nurse cells, and the oocyte. Genes defined by a common RNA localization share distinct gene features and differ in expression level, 3'UTR length and sequence conservation from unlocalized mRNAs. Comparison of mRNA localizations in different contexts revealed that localization of individual mRNAs changes over time in the oocyte and between ovarian and embryonic cell types. This genome scale image-based resource (Dresden Ovary Table, DOT, http://tomancak-srv1.mpi-cbg.de/DOT/main.html) enables the transition from mechanistic dissection of singular mRNA localization events towards global understanding of how mRNAs transcribed in the nucleus distribute in cells.

Show MeSH
Related in: MedlinePlus