Limits...
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.

Show MeSH

Related in: MedlinePlus

Cytoplasmic but not nuclear mRNA localization requires the cytoskeleton.(A) Localization of anterior and posterior mRNAs is lost upon microtubule depolymerization by colchicine. Shown are the anterior mRNAs fs(1)K10 and milt (examples for diffuse-anterior and tight-anterior localization) and the and posterior mRNA vkg and zpg (examples for diffuse-posterior and tight-posterior). The appearance of the ubiquitous mRNA msl-2 is unchanged in colchicine treated egg-chambers. For details see Supplementary file 8. (B) Summary of the quality of all anterior and posterior mRNA distributions tested in colchicine treated egg-chambers (round aggregates, tiny aggregates, dispersed and diffuse aggregates). Diffuse aggregates were observed for those mRNAs that in wild type egg-chambers showed a diffuse posterior enrichment (e.g., Figure 1B’: fs(1)N). (C) mRNA localization in proximity to the nucleus is lost in colchicine treated egg-chambers (Scp2), RNAs localized partially nuclear and partially perinuclear loose the cytoplasmic localization (CG11076) while strictly nuclear RNAs are unaffected by microtubule depolymerization (rhi). (A,C) FISH experiments showing the RNA in green; DNA (labelled with DAPI) is shown in magenta (A) or blue (C), the nuclear membrane is stained with WGA dye shown in red (C). Scale bar 30 μm.DOI:http://dx.doi.org/10.7554/eLife.05003.012
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4384636&req=5

fig3s4: Cytoplasmic but not nuclear mRNA localization requires the cytoskeleton.(A) Localization of anterior and posterior mRNAs is lost upon microtubule depolymerization by colchicine. Shown are the anterior mRNAs fs(1)K10 and milt (examples for diffuse-anterior and tight-anterior localization) and the and posterior mRNA vkg and zpg (examples for diffuse-posterior and tight-posterior). The appearance of the ubiquitous mRNA msl-2 is unchanged in colchicine treated egg-chambers. For details see Supplementary file 8. (B) Summary of the quality of all anterior and posterior mRNA distributions tested in colchicine treated egg-chambers (round aggregates, tiny aggregates, dispersed and diffuse aggregates). Diffuse aggregates were observed for those mRNAs that in wild type egg-chambers showed a diffuse posterior enrichment (e.g., Figure 1B’: fs(1)N). (C) mRNA localization in proximity to the nucleus is lost in colchicine treated egg-chambers (Scp2), RNAs localized partially nuclear and partially perinuclear loose the cytoplasmic localization (CG11076) while strictly nuclear RNAs are unaffected by microtubule depolymerization (rhi). (A,C) FISH experiments showing the RNA in green; DNA (labelled with DAPI) is shown in magenta (A) or blue (C), the nuclear membrane is stained with WGA dye shown in red (C). Scale bar 30 μm.DOI:http://dx.doi.org/10.7554/eLife.05003.012

Mentions: The posterior gene set shows clearly distinct functional and gene architectural features compared to all the other categories. We therefore decided to investigate whether the cytoplasmic localization of the novel candidate mRNAs depends on the known components of RNA localization machinery in the oocyte. First, we probed the dependency of mRNA localization on the microtubule cytoskeleton. Transport of known mRNAs towards the anterior and the posterior pole of the oocyte requires an intact microtubule cytoskeleton (reviewed in Steinhauer and Kalderon, 2006). We observed that the localization of all new anterior and posterior candidate mRNAs is lost in colchicine-treated egg-chambers, while ubiquitously distributed mRNAs or RNA foci in the nucleoplasm, that lacks a microtubule cytoskeleton, were unaffected by the colchicine treatment (Figure 3—figure supplement 4A–C, Supplementary file 8).


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)

Cytoplasmic but not nuclear mRNA localization requires the cytoskeleton.(A) Localization of anterior and posterior mRNAs is lost upon microtubule depolymerization by colchicine. Shown are the anterior mRNAs fs(1)K10 and milt (examples for diffuse-anterior and tight-anterior localization) and the and posterior mRNA vkg and zpg (examples for diffuse-posterior and tight-posterior). The appearance of the ubiquitous mRNA msl-2 is unchanged in colchicine treated egg-chambers. For details see Supplementary file 8. (B) Summary of the quality of all anterior and posterior mRNA distributions tested in colchicine treated egg-chambers (round aggregates, tiny aggregates, dispersed and diffuse aggregates). Diffuse aggregates were observed for those mRNAs that in wild type egg-chambers showed a diffuse posterior enrichment (e.g., Figure 1B’: fs(1)N). (C) mRNA localization in proximity to the nucleus is lost in colchicine treated egg-chambers (Scp2), RNAs localized partially nuclear and partially perinuclear loose the cytoplasmic localization (CG11076) while strictly nuclear RNAs are unaffected by microtubule depolymerization (rhi). (A,C) FISH experiments showing the RNA in green; DNA (labelled with DAPI) is shown in magenta (A) or blue (C), the nuclear membrane is stained with WGA dye shown in red (C). Scale bar 30 μm.DOI:http://dx.doi.org/10.7554/eLife.05003.012
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4384636&req=5

fig3s4: Cytoplasmic but not nuclear mRNA localization requires the cytoskeleton.(A) Localization of anterior and posterior mRNAs is lost upon microtubule depolymerization by colchicine. Shown are the anterior mRNAs fs(1)K10 and milt (examples for diffuse-anterior and tight-anterior localization) and the and posterior mRNA vkg and zpg (examples for diffuse-posterior and tight-posterior). The appearance of the ubiquitous mRNA msl-2 is unchanged in colchicine treated egg-chambers. For details see Supplementary file 8. (B) Summary of the quality of all anterior and posterior mRNA distributions tested in colchicine treated egg-chambers (round aggregates, tiny aggregates, dispersed and diffuse aggregates). Diffuse aggregates were observed for those mRNAs that in wild type egg-chambers showed a diffuse posterior enrichment (e.g., Figure 1B’: fs(1)N). (C) mRNA localization in proximity to the nucleus is lost in colchicine treated egg-chambers (Scp2), RNAs localized partially nuclear and partially perinuclear loose the cytoplasmic localization (CG11076) while strictly nuclear RNAs are unaffected by microtubule depolymerization (rhi). (A,C) FISH experiments showing the RNA in green; DNA (labelled with DAPI) is shown in magenta (A) or blue (C), the nuclear membrane is stained with WGA dye shown in red (C). Scale bar 30 μm.DOI:http://dx.doi.org/10.7554/eLife.05003.012
Mentions: The posterior gene set shows clearly distinct functional and gene architectural features compared to all the other categories. We therefore decided to investigate whether the cytoplasmic localization of the novel candidate mRNAs depends on the known components of RNA localization machinery in the oocyte. First, we probed the dependency of mRNA localization on the microtubule cytoskeleton. Transport of known mRNAs towards the anterior and the posterior pole of the oocyte requires an intact microtubule cytoskeleton (reviewed in Steinhauer and Kalderon, 2006). We observed that the localization of all new anterior and posterior candidate mRNAs is lost in colchicine-treated egg-chambers, while ubiquitously distributed mRNAs or RNA foci in the nucleoplasm, that lacks a microtubule cytoskeleton, were unaffected by the colchicine treatment (Figure 3—figure supplement 4A–C, Supplementary file 8).

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