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Drosophila Ge-1 promotes P body formation and oskar mRNA localization.

Fan SJ, Marchand V, Ephrussi A - PLoS ONE (2011)

Bottom Line: Several P body components are involved in osk mRNA localization and translational repression, suggesting a link between P bodies and osk RNPs.In cultured mammalian cells, Ge-1 protein is required for P body formation.Our findings suggest an important role of dGe-1 in optimization of the osk mRNA localization process required for patterning the Drosophila embryo.

View Article: PubMed Central - PubMed

Affiliation: Developmental Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany.

ABSTRACT
mRNA localization coupled with translational control is a widespread and conserved strategy that allows the localized production of proteins within eukaryotic cells. In Drosophila, oskar (osk) mRNA localization and translation at the posterior pole of the oocyte are essential for proper patterning of the embryo. Several P body components are involved in osk mRNA localization and translational repression, suggesting a link between P bodies and osk RNPs. In cultured mammalian cells, Ge-1 protein is required for P body formation. Combining genetic, biochemical and immunohistochemical approaches, we show that, in vivo, Drosophila Ge-1 (dGe-1) is an essential gene encoding a P body component that promotes formation of these structures in the germline. dGe-1 partially colocalizes with osk mRNA and is required for osk RNP integrity. Our analysis reveals that although under normal conditions dGe-1 function is not essential for osk mRNA localization, it becomes critical when other components of the localization machinery, such as staufen, Drosophila decapping protein 1 and barentsz are limiting. Our findings suggest an important role of dGe-1 in optimization of the osk mRNA localization process required for patterning the Drosophila embryo.

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dGe-1 partially colocalizes with osk mRNA.(A–C) Immunodetection of dGe-1 protein in wt (A, B) and dGe-1Δ5 GLC (C) ovaries using a rabbit anti-dGe-1 antibody. (A, B) In wt ovaries at S9 (A) or S10 (B), dGe-1 protein is enriched at the oocyte posterior pole. (C) The posterior dGe-1 signal observed in wt oocytes (B) is greatly reduced in dGe-1Δ5 GLC oocytes. (D–F) In a wt egg-chamber at late S9, dGe-1 enriched at the posterior pole of the oocyte (green, D) and partially colocalizes with Stau protein (red, E). The overlay of the two antibody signals is shown in (F). (G–I) Colocalization of dGe-1 protein (green, G) and osk mRNA (red, H) in wt S9 oocytes expressing oskMS2 mRNA and MCP-RFP protein (red, H), to which the RNA is tethered; at S9 oskMS2 mRNA distribution reflects that of endogenous osk mRNA [33]. Overlay of dGe-1 and oskMS2 mRNA signals shown in (I). (J–K) dGe-1 protein staining in wt and stauD3 oocytes. (J, J′) dGe-1 protein is enriched at the posterior pole in wt oocytes. (K and K′) In stauD3 mutant oocytes, which fail to localize osk mRNA, the amount of dGe-1 at the posterior is dramatically reduced. (L–N) Double immunostaining of dGe-1 and Stau proteins in a nutritionally restricted wt egg-chamber. In starved wt oocytes, dGe-1 protein (green, L) is present in large particles, where it colocalizes with Stau protein (red, M). Overlay of dGe-1 and Stau stainings (N). (O–Q) Distribution of dGe-1 protein and osk mRNA in starved wt egg-chambers expressing oskBoxB mRNA and λN-GFP protein, to which the RNA is tethered. In nutritionally restricted wt oocytes, dGe-1 (red, O) is present in large particles, where it partially colocalizes with oskBoxB mRNA (green, P). Overlay of dGe-1 and osk mRNA signals shown in (Q). Bar, 50 µm.
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pone-0020612-g004: dGe-1 partially colocalizes with osk mRNA.(A–C) Immunodetection of dGe-1 protein in wt (A, B) and dGe-1Δ5 GLC (C) ovaries using a rabbit anti-dGe-1 antibody. (A, B) In wt ovaries at S9 (A) or S10 (B), dGe-1 protein is enriched at the oocyte posterior pole. (C) The posterior dGe-1 signal observed in wt oocytes (B) is greatly reduced in dGe-1Δ5 GLC oocytes. (D–F) In a wt egg-chamber at late S9, dGe-1 enriched at the posterior pole of the oocyte (green, D) and partially colocalizes with Stau protein (red, E). The overlay of the two antibody signals is shown in (F). (G–I) Colocalization of dGe-1 protein (green, G) and osk mRNA (red, H) in wt S9 oocytes expressing oskMS2 mRNA and MCP-RFP protein (red, H), to which the RNA is tethered; at S9 oskMS2 mRNA distribution reflects that of endogenous osk mRNA [33]. Overlay of dGe-1 and oskMS2 mRNA signals shown in (I). (J–K) dGe-1 protein staining in wt and stauD3 oocytes. (J, J′) dGe-1 protein is enriched at the posterior pole in wt oocytes. (K and K′) In stauD3 mutant oocytes, which fail to localize osk mRNA, the amount of dGe-1 at the posterior is dramatically reduced. (L–N) Double immunostaining of dGe-1 and Stau proteins in a nutritionally restricted wt egg-chamber. In starved wt oocytes, dGe-1 protein (green, L) is present in large particles, where it colocalizes with Stau protein (red, M). Overlay of dGe-1 and Stau stainings (N). (O–Q) Distribution of dGe-1 protein and osk mRNA in starved wt egg-chambers expressing oskBoxB mRNA and λN-GFP protein, to which the RNA is tethered. In nutritionally restricted wt oocytes, dGe-1 (red, O) is present in large particles, where it partially colocalizes with oskBoxB mRNA (green, P). Overlay of dGe-1 and osk mRNA signals shown in (Q). Bar, 50 µm.

Mentions: In addition to its punctate distribution in the nurse cells, dGe-1 protein is also observed in a crescent at the oocyte posterior during mid-oogenesis (Figure 4A–B). This staining is specific, as it is dramatically reduced in dGe-1Δ5 mutant oocytes (Figure 4C). Moreover, the posterior crescent of dGe-1 overlaps with that of Stau, an osk mRNA associated protein, and with osk mRNA (Figure 4D–I), suggesting that dGe-1 protein is a component of osk RNPs.


Drosophila Ge-1 promotes P body formation and oskar mRNA localization.

Fan SJ, Marchand V, Ephrussi A - PLoS ONE (2011)

dGe-1 partially colocalizes with osk mRNA.(A–C) Immunodetection of dGe-1 protein in wt (A, B) and dGe-1Δ5 GLC (C) ovaries using a rabbit anti-dGe-1 antibody. (A, B) In wt ovaries at S9 (A) or S10 (B), dGe-1 protein is enriched at the oocyte posterior pole. (C) The posterior dGe-1 signal observed in wt oocytes (B) is greatly reduced in dGe-1Δ5 GLC oocytes. (D–F) In a wt egg-chamber at late S9, dGe-1 enriched at the posterior pole of the oocyte (green, D) and partially colocalizes with Stau protein (red, E). The overlay of the two antibody signals is shown in (F). (G–I) Colocalization of dGe-1 protein (green, G) and osk mRNA (red, H) in wt S9 oocytes expressing oskMS2 mRNA and MCP-RFP protein (red, H), to which the RNA is tethered; at S9 oskMS2 mRNA distribution reflects that of endogenous osk mRNA [33]. Overlay of dGe-1 and oskMS2 mRNA signals shown in (I). (J–K) dGe-1 protein staining in wt and stauD3 oocytes. (J, J′) dGe-1 protein is enriched at the posterior pole in wt oocytes. (K and K′) In stauD3 mutant oocytes, which fail to localize osk mRNA, the amount of dGe-1 at the posterior is dramatically reduced. (L–N) Double immunostaining of dGe-1 and Stau proteins in a nutritionally restricted wt egg-chamber. In starved wt oocytes, dGe-1 protein (green, L) is present in large particles, where it colocalizes with Stau protein (red, M). Overlay of dGe-1 and Stau stainings (N). (O–Q) Distribution of dGe-1 protein and osk mRNA in starved wt egg-chambers expressing oskBoxB mRNA and λN-GFP protein, to which the RNA is tethered. In nutritionally restricted wt oocytes, dGe-1 (red, O) is present in large particles, where it partially colocalizes with oskBoxB mRNA (green, P). Overlay of dGe-1 and osk mRNA signals shown in (Q). Bar, 50 µm.
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Related In: Results  -  Collection

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pone-0020612-g004: dGe-1 partially colocalizes with osk mRNA.(A–C) Immunodetection of dGe-1 protein in wt (A, B) and dGe-1Δ5 GLC (C) ovaries using a rabbit anti-dGe-1 antibody. (A, B) In wt ovaries at S9 (A) or S10 (B), dGe-1 protein is enriched at the oocyte posterior pole. (C) The posterior dGe-1 signal observed in wt oocytes (B) is greatly reduced in dGe-1Δ5 GLC oocytes. (D–F) In a wt egg-chamber at late S9, dGe-1 enriched at the posterior pole of the oocyte (green, D) and partially colocalizes with Stau protein (red, E). The overlay of the two antibody signals is shown in (F). (G–I) Colocalization of dGe-1 protein (green, G) and osk mRNA (red, H) in wt S9 oocytes expressing oskMS2 mRNA and MCP-RFP protein (red, H), to which the RNA is tethered; at S9 oskMS2 mRNA distribution reflects that of endogenous osk mRNA [33]. Overlay of dGe-1 and oskMS2 mRNA signals shown in (I). (J–K) dGe-1 protein staining in wt and stauD3 oocytes. (J, J′) dGe-1 protein is enriched at the posterior pole in wt oocytes. (K and K′) In stauD3 mutant oocytes, which fail to localize osk mRNA, the amount of dGe-1 at the posterior is dramatically reduced. (L–N) Double immunostaining of dGe-1 and Stau proteins in a nutritionally restricted wt egg-chamber. In starved wt oocytes, dGe-1 protein (green, L) is present in large particles, where it colocalizes with Stau protein (red, M). Overlay of dGe-1 and Stau stainings (N). (O–Q) Distribution of dGe-1 protein and osk mRNA in starved wt egg-chambers expressing oskBoxB mRNA and λN-GFP protein, to which the RNA is tethered. In nutritionally restricted wt oocytes, dGe-1 (red, O) is present in large particles, where it partially colocalizes with oskBoxB mRNA (green, P). Overlay of dGe-1 and osk mRNA signals shown in (Q). Bar, 50 µm.
Mentions: In addition to its punctate distribution in the nurse cells, dGe-1 protein is also observed in a crescent at the oocyte posterior during mid-oogenesis (Figure 4A–B). This staining is specific, as it is dramatically reduced in dGe-1Δ5 mutant oocytes (Figure 4C). Moreover, the posterior crescent of dGe-1 overlaps with that of Stau, an osk mRNA associated protein, and with osk mRNA (Figure 4D–I), suggesting that dGe-1 protein is a component of osk RNPs.

Bottom Line: Several P body components are involved in osk mRNA localization and translational repression, suggesting a link between P bodies and osk RNPs.In cultured mammalian cells, Ge-1 protein is required for P body formation.Our findings suggest an important role of dGe-1 in optimization of the osk mRNA localization process required for patterning the Drosophila embryo.

View Article: PubMed Central - PubMed

Affiliation: Developmental Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany.

ABSTRACT
mRNA localization coupled with translational control is a widespread and conserved strategy that allows the localized production of proteins within eukaryotic cells. In Drosophila, oskar (osk) mRNA localization and translation at the posterior pole of the oocyte are essential for proper patterning of the embryo. Several P body components are involved in osk mRNA localization and translational repression, suggesting a link between P bodies and osk RNPs. In cultured mammalian cells, Ge-1 protein is required for P body formation. Combining genetic, biochemical and immunohistochemical approaches, we show that, in vivo, Drosophila Ge-1 (dGe-1) is an essential gene encoding a P body component that promotes formation of these structures in the germline. dGe-1 partially colocalizes with osk mRNA and is required for osk RNP integrity. Our analysis reveals that although under normal conditions dGe-1 function is not essential for osk mRNA localization, it becomes critical when other components of the localization machinery, such as staufen, Drosophila decapping protein 1 and barentsz are limiting. Our findings suggest an important role of dGe-1 in optimization of the osk mRNA localization process required for patterning the Drosophila embryo.

Show MeSH
Related in: MedlinePlus