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  1. Article ; Online: Basic Techniques in Drosophila Ovary Preparation.

    Thompson, Letitia / Randolph, Kristen / Norvell, Amanda

    Methods in molecular biology (Clifton, N.J.)

    2015  Volume 1328, Page(s) 21–28

    Abstract: Drosophila melanogaster oogenesis has emerged as an excellent model system to study multiple aspects of eukaryotic cell biology. Ovarian tissue can easily be isolated and analyzed through microscopy or biochemical and molecular biology techniques. Here ... ...

    Abstract Drosophila melanogaster oogenesis has emerged as an excellent model system to study multiple aspects of eukaryotic cell biology. Ovarian tissue can easily be isolated and analyzed through microscopy or biochemical and molecular biology techniques. Here we describe the isolation of ovarian tissues, techniques to enrich for egg chambers at distinct developmental stages, preparation of protein and nucleic acid extracts, and preparation for microscopic analysis of fixed tissues.
    MeSH term(s) Animals ; Drosophila melanogaster/genetics ; Drosophila melanogaster/growth & development ; Female ; Molecular Biology/methods ; Oocytes/growth & development ; Oogenesis ; Ovary
    Language English
    Publishing date 2015
    Publishing country United States
    Document type Journal Article
    ISSN 1940-6029
    ISSN (online) 1940-6029
    DOI 10.1007/978-1-4939-2851-4_2
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  2. Article ; Online: Wispy and Orb cooperate in the cytoplasmic polyadenylation of localized gurken mRNA.

    Norvell, Amanda / Wong, Jason / Randolph, Kristen / Thompson, Letitia

    Developmental dynamics : an official publication of the American Association of Anatomists

    2015  Volume 244, Issue 10, Page(s) 1276–1285

    Abstract: Background: In Drosophila, the dorsal-ventral (D-V) axis of the oocyte is dependent on Gurken (Grk) protein distribution. This is achieved through the cytoplasmic localization of grk mRNA and regulation of its translation. During mid-late stages of ... ...

    Abstract Background: In Drosophila, the dorsal-ventral (D-V) axis of the oocyte is dependent on Gurken (Grk) protein distribution. This is achieved through the cytoplasmic localization of grk mRNA and regulation of its translation. During mid-late stages of oogenesis, grk mRNA and protein are localized to the dorsal-anterior of the oocyte, while unlocalized grk transcripts are translationally silenced. As females carrying mutations in the gene encoding the CPEB protein Orb lay ventralized eggs due to insufficient Grk levels, it seemed likely that cytoplasmic polyadenylation of grk transcripts may play a role in their translational regulation.
    Results: We have found that grk is polyadenylated throughout oogenesis, with poly(A) tails of approximately 30-50 A residues. Hyperadenylated grk transcripts, with poly(A) tails of 50-90 As, are detected in late stage egg chambers, but they fail to accumulate in oocytes deficient in Orb or the poly(A) polymerase Wispy (Wisp). wisp females also lay weakly ventralized eggs, demonstrating that they produce inadequate amounts of Grk. Finally, unlocalized grk transcripts are also not appropriately hyperadenylated.
    Conclusions: Localized cytoplasmic polyadenylation of grk mRNA by Wisp and Orb is necessary to achieve appropriate Grk protein accumulation in the D/A corner of the oocyte during mid to late oogenesis.
    MeSH term(s) Animals ; Body Patterning ; Drosophila Proteins/genetics ; Drosophila Proteins/metabolism ; Drosophila melanogaster/genetics ; Drosophila melanogaster/metabolism ; Female ; Male ; Oogenesis ; Polyadenylation ; Polynucleotide Adenylyltransferase/metabolism ; RNA-Binding Proteins/genetics ; RNA-Binding Proteins/metabolism ; Transforming Growth Factor alpha/genetics
    Chemical Substances Drosophila Proteins ; RNA-Binding Proteins ; Transforming Growth Factor alpha ; grk protein, Drosophila ; orb protein, Drosophila ; Polynucleotide Adenylyltransferase (EC 2.7.7.19) ; wisp protein, Drosophila (EC 2.7.7.19)
    Language English
    Publishing date 2015-08-26
    Publishing country United States
    Document type Journal Article ; Research Support, U.S. Gov't, Non-P.H.S.
    ZDB-ID 1102541-4
    ISSN 1097-0177 ; 1058-8388
    ISSN (online) 1097-0177
    ISSN 1058-8388
    DOI 10.1002/dvdy.24311
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  3. Article ; Online: Cell biology. Rise of the rival.

    Norvell, Amanda / McMahon, Steven B

    Science (New York, N.Y.)

    2010  Volume 327, Issue 5968, Page(s) 964–965

    MeSH term(s) Acetyl Coenzyme A/metabolism ; Acetylation ; Acetylesterase/metabolism ; Bacterial Proteins/metabolism ; Enzymes/metabolism ; Hepatocytes/enzymology ; Hepatocytes/metabolism ; Humans ; Lysine/metabolism ; Metabolic Networks and Pathways ; Phosphorylation ; Proteins/metabolism ; Salmonella enterica/enzymology ; Salmonella enterica/metabolism
    Chemical Substances Bacterial Proteins ; Enzymes ; Proteins ; Acetyl Coenzyme A (72-89-9) ; Acetylesterase (EC 3.1.1.6) ; Lysine (K3Z4F929H6)
    Language English
    Publishing date 2010-02-19
    Publishing country United States
    Document type Comment ; Journal Article
    ZDB-ID 128410-1
    ISSN 1095-9203 ; 0036-8075
    ISSN (online) 1095-9203
    ISSN 0036-8075
    DOI 10.1126/science.1187159
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  4. Article: Squid is required for efficient posterior localization of oskar mRNA during Drosophila oogenesis.

    Norvell, Amanda / Debec, Alain / Finch, Daniel / Gibson, Lisa / Thoma, Brandi

    Development genes and evolution

    2005  Volume 215, Issue 7, Page(s) 340–349

    Abstract: The nuclear-cytoplasmic shuttling heterogeneous nuclear RNA-binding protein (hnRNP) Squid (Sqd) is required during Drosophila melanogaster oogenesis, where it plays a critical role in the regulation of the TGFalpha-like molecule Gurken (Grk). Three Sqd ... ...

    Abstract The nuclear-cytoplasmic shuttling heterogeneous nuclear RNA-binding protein (hnRNP) Squid (Sqd) is required during Drosophila melanogaster oogenesis, where it plays a critical role in the regulation of the TGFalpha-like molecule Gurken (Grk). Three Sqd isoforms have been described, SqdA, S and B, and two of these, SqdA and SqdS, differentially function in grk mRNA nuclear export, cytoplasmic transport and translational control during oogenesis. Here, we report that Sqd is also required for the regulation of oskar (osk) mRNA, functioning in the cytoplasmic localization of the osk transcript. In oocytes from sqd females, osk mRNA is not efficiently localized to the posterior pole, but rather accumulates at the anterior cortex. Furthermore, anterior patterning defects observed in embryos from sqd females expressing only the SqdS protein isoform suggest that Sqd may also play a role in the translational regulation of the mislocalized osk mRNA. These findings provide additional support for models of mRNA regulation in which cytoplasmic events, such as localization and translational regulation, are coupled. These results also place Sqd among an emerging class of proteins, including such other members as Bruno (Bru) and Hrb27C/Hrp48, which function in multiple aspects of both grk and osk mRNA regulation during Drosophila oogenesis.
    MeSH term(s) 3' Untranslated Regions ; Animals ; Body Patterning ; Cross-Linking Reagents/pharmacology ; Drosophila/anatomy & histology ; Drosophila/physiology ; Drosophila Proteins/genetics ; Drosophila Proteins/metabolism ; Female ; Green Fluorescent Proteins/metabolism ; Immunohistochemistry ; In Situ Hybridization ; Mutation ; Oocytes/physiology ; Oogenesis/physiology ; Ovary/cytology ; Ovary/physiology ; Ovary/radiation effects ; Protein Binding ; RNA, Messenger/genetics ; RNA, Messenger/metabolism ; RNA-Binding Proteins/genetics ; RNA-Binding Proteins/metabolism ; Recombinant Fusion Proteins/metabolism ; Ultraviolet Rays
    Chemical Substances 3' Untranslated Regions ; Cross-Linking Reagents ; Drosophila Proteins ; RNA, Messenger ; RNA-Binding Proteins ; Recombinant Fusion Proteins ; osk protein, Drosophila ; sqd protein, Drosophila ; Green Fluorescent Proteins (147336-22-9)
    Language English
    Publishing date 2005-03-25
    Publishing country Germany
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 1327962-2
    ISSN 1432-041X ; 0949-944X
    ISSN (online) 1432-041X
    ISSN 0949-944X
    DOI 10.1007/s00427-005-0480-2
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