An Interaction Type of Genetic Screen Reveals a Role of the Rab11 Gene in oskar mRNA Localization in the Developing Drosophila melanogaster Oocyte

Abstract Abdomen and germ cell development of Drosophila melanogaster embryo requires proper localization of oskar mRNA to the posterior pole of the developing oocyte. oskar mRNA localization depends on complex cell biological events like cell-cell communication, dynamic rearrangement of the microtubule network, and function of the actin cytoskeleton of the oocyte. To investigate the cellular mechanisms involved, we developed a novel interaction type of genetic screen by which we isolated 14 dominant enhancers of a sensitized genetic background composed of mutations in oskar and in TropomyosinII, an actin binding protein. Here we describe the detailed analysis of two allelic modifiers that identify Drosophila Rab11, a gene encoding small monomeric GTPase. We demonstrate that mutation of the Rab11 gene, involved in various vesicle transport processes, results in ectopic localization of oskar mRNA, whereas localization of gurken and bicoid mRNAs and signaling between the oocyte and the somatic follicle cells are unaffected. We show that the ectopic oskar mRNA localization in the Rab11 mutants is a consequence of an abnormally polarized oocyte microtubule cytoskeleton. Our results indicate that the internal membranous structures play an important role in the microtubule organization in the Drosophila oocyte and, thus, in oskar RNA localization.

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Oskar anchoring restricts pole plasm formation to the posterior of the Drosophila oocyte

Development ◽

10.1242/dev.129.15.3705 ◽

2002 ◽

Vol 129 (15) ◽

pp. 3705-3714 ◽

Cited By ~ 14

Author(s):

Nathalie F. Vanzo ◽

Anne Ephrussi

Keyword(s):

Drosophila Melanogaster ◽

Translational Control ◽

Cell Formation ◽

Positional Information ◽

Mrna Localization ◽

Posterior Pole ◽

Tight Coupling ◽

Anteroposterior Patterning ◽

Pole Cell ◽

Pole Plasm

Localization of the maternal determinant Oskar at the posterior pole of Drosophila melanogaster oocyte provides the positional information for pole plasm formation. Spatial control of Oskar expression is achieved through the tight coupling of mRNA localization to translational control, such that only posterior-localized oskar mRNA is translated, producing the two Oskar isoforms Long Osk and Short Osk. We present evidence that this coupling is not sufficient to restrict Oskar to the posterior pole of the oocyte. We show that Long Osk anchors both oskar mRNA and Short Osk, the isoform active in pole plasm assembly, at the posterior pole. In the absence of anchoring by Long Osk, Short Osk disperses into the bulk cytoplasm during late oogenesis, impairing pole cell formation in the embryo. In addition, the pool of untethered Short Osk causes anteroposterior patterning defects, owing to the dispersion of pole plasm and its abdomen-inducing activity throughout the oocyte. We show that the N-terminal extension of Long Osk is necessary but not sufficient for posterior anchoring, arguing for multiple docking elements in Oskar. This study reveals cortical anchoring of the posterior determinant Oskar as a crucial step in pole plasm assembly and restriction, required for proper development of Drosophila melanogaster.

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The Spectraplakin Short Stop Is an Actin–Microtubule Cross-Linker That Contributes to Organization of the Microtubule Network

Molecular Biology of the Cell ◽

10.1091/mbc.e10-01-0011 ◽

2010 ◽

Vol 21 (10) ◽

pp. 1714-1724 ◽

Cited By ~ 77

Author(s):

Derek A. Applewhite ◽

Kyle D. Grode ◽

Darby Keller ◽

Alireza Dehghani Zadeh ◽

Kevin C. Slep ◽

...

Keyword(s):

Cross Talk ◽

Function Analysis ◽

Cytoskeletal Proteins ◽

Actin Binding ◽

Cross Linking ◽

Cell Periphery ◽

Cellular Functions ◽

Microtubule Organization ◽

Microtubule Network ◽

Cross Linker

The dynamics of actin and microtubules are coordinated in a variety of cellular and morphogenetic processes; however, little is known about the molecules mediating this cytoskeletal cross-talk. We are studying Short stop (Shot), the sole Drosophila spectraplakin, as a model actin–microtubule cross-linking protein. Spectraplakins are an ancient family of giant cytoskeletal proteins that are essential for a diverse set of cellular functions; yet, we know little about the dynamics of spectraplakins and how they bridge actin filaments and microtubules. In this study we describe the intracellular dynamics of Shot and a structure–function analysis of its role as a cytoskeletal cross-linker. We find that Shot interacts with microtubules using two different mechanisms. In the cell interior, Shot binds growing plus ends through an interaction with EB1. In the cell periphery, Shot associates with the microtubule lattice via its GAS2 domain, and this pool of Shot is actively engaged as a cross-linker via its NH2-terminal actin-binding calponin homology domains. This cross-linking maintains microtubule organization by resisting forces that produce lateral microtubule movements in the cytoplasm. Our results provide the first description of the dynamics of these important proteins and provide key insight about how they function during cytoskeletal cross-talk.

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Drosophila virilis oskar transgenes direct body patterning but not pole cell formation or maintenance of mRNA localization in D. melanogaster

Development ◽

10.1242/dev.120.7.2027 ◽

1994 ◽

Vol 120 (7) ◽

pp. 2027-2037 ◽

Cited By ~ 2

Author(s):

P.J. Webster ◽

J. Suen ◽

P.M. Macdonald

Keyword(s):

Drosophila Melanogaster ◽

Germ Cell ◽

Cell Formation ◽

Mrna Localization ◽

Early Embryo ◽

Posterior Pole ◽

Early Embryogenesis ◽

Drosophila Virilis ◽

Pole Cell ◽

Body Patterning

The Drosophila melanogaster gene oskar is required for both posterior body patterning and germline formation in the early embryo; precisely how oskar functions is unknown. The oskar transcript is localized to the posterior pole of the developing oocyte, and oskar mRNA and protein are maintained at the pole through early embryogenesis. The posterior maintenance of oskar mRNA is dependent upon the presence of oskar protein. We have cloned and characterized the Drosophila virilis oskar homologue, virosk, and examined its activity as a transgene in Drosophila melanogaster flies. We find that the cis-acting mRNA localization signals are conserved, although the virosk transcript also transiently accumulates at novel intermediate sites. The virosk protein, however, shows substantial differences from oskar: while virosk is able to rescue body patterning in a D. melanogaster oskar- background, it is impaired in both mRNA maintenance and pole cell formation. Furthermore, virosk induces a dominant maternal-effect lethality when introduced into a wild-type background, and interferes with the posterior maintenance of the endogenous oskar transcript in early embryogenesis. Our data suggest that virosk protein is unable to anchor at the posterior pole of the early embryo; this defect could account for all of the characteristics of virosk mentioned above. Our observations support a model in which oskar protein functions both by nucleating the factors necessary for the activation of the posterior body patterning determinant and the germ cell determinant, and by anchoring these factors to the posterior pole of the embryo. While the posterior body patterning determinant need not be correctly localized to provide body patterning activity, the germ cell determinant may need to be highly concentrated adjacent to the cortex in order to direct pole cell formation.

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Wolbachia Influences the Maternal Transmission of the gypsy Endogenous Retrovirus in Drosophila melanogaster

mBio ◽

10.1128/mbio.01529-14 ◽

2014 ◽

Vol 5 (5) ◽

Cited By ~ 15

Author(s):

Franck Touret ◽

François Guiguen ◽

Christophe Terzian

Keyword(s):

Drosophila Melanogaster ◽

Antiviral Activity ◽

Rna Viruses ◽

Natural Populations ◽

Endogenous Retrovirus ◽

Posterior Pole ◽

Endogenous Retroviruses ◽

Follicle Cells ◽

Maternal Transmission ◽

Content Type

ABSTRACT The endosymbiotic bacteria of the genus Wolbachia are present in most insects and are maternally transmitted through the germline. Moreover, these intracellular bacteria exert antiviral activity against insect RNA viruses, as in Drosophila melanogaster, which could explain the prevalence of Wolbachia bacteria in natural populations. Wolbachia is maternally transmitted in D. melanogaster through a mechanism that involves distribution at the posterior pole of mature oocytes and then incorporation into the pole cells of the embryos. In parallel, maternal transmission of several endogenous retroviruses is well documented in D. melanogaster. Notably, gypsy retrovirus is expressed in permissive follicle cells and transferred to the oocyte and then to the offspring by integrating into their genomes. Here, we show that the presence of Wolbachia wMel reduces the rate of gypsy insertion into the ovo gene. However, the presence of Wolbachia does not modify the expression levels of gypsy RNA and envelope glycoprotein from either permissive or restrictive ovaries. Moreover, Wolbachia affects the pattern of distribution of the retroviral particles and the gypsy envelope protein in permissive follicle cells. Altogether, our results enlarge the knowledge of the antiviral activity of Wolbachia to include reducing the maternal transmission of endogenous retroviruses in D. melanogaster. IMPORTANCE Animals have established complex relationships with bacteria and viruses that spread horizontally among individuals or are vertically transmitted, i.e., from parents to offspring. It is well established that members of the genus Wolbachia, maternally inherited symbiotic bacteria present mainly in arthropods, reduce the replication of several RNA viruses transmitted horizontally. Here, we demonstrate for the first time that Wolbachia diminishes the maternal transmission of gypsy, an endogenous retrovirus in Drosophila melanogaster. We hypothesize that gypsy cannot efficiently integrate into the germ cells of offspring during embryonic development in the presence of Wolbachia because both are competitors for localization to the posterior pole of the egg. More generally, it would be of interest to analyze the influence of Wolbachia on vertically transmitted exogenous viruses, such as some arboviruses.

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A Genetic Screen Reveals an Unexpected Role for Yorkie Signaling in JAK/STAT-Dependent Hematopoietic Malignancies in Drosophila melanogaster

G3 Genes|Genome|Genetics ◽

10.1534/g3.117.044172 ◽

2017 ◽

Vol 7 (8) ◽

pp. 2427-2438 ◽

Cited By ~ 12

Author(s):

Abigail M. Anderson ◽

Alessandro A. Bailetti ◽

Elizabeth Rodkin ◽

Atish De ◽

Erika A. Bach

Keyword(s):

Drosophila Melanogaster ◽

Genetic Screen ◽

Hematopoietic Malignancies

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Intercellular bridges in ovarian follicle cells of Drosophila melanogaster

Cell and Tissue Research ◽

10.1007/bf00224931 ◽

1978 ◽

Vol 186 (3) ◽

Cited By ~ 21

Author(s):

Franco Giorgi

Keyword(s):

Drosophila Melanogaster ◽

Ovarian Follicle ◽

Follicle Cells ◽

Intercellular Bridges

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Pvr expression regulators in equilibrium signal control and maintenance of Drosophila blood progenitors

eLife ◽

10.7554/elife.03626 ◽

2014 ◽

Vol 3 ◽

Cited By ~ 37

Author(s):

Bama Charan Mondal ◽

Jiwon Shim ◽

Cory J Evans ◽

Utpal Banerjee

Keyword(s):

Drosophila Melanogaster ◽

Blood Cells ◽

Lymph Gland ◽

Genetic Screen ◽

Signal Control ◽

Vegf Receptor ◽

Interacting Protein ◽

Hematopoietic Development ◽

New Genes ◽

Progenitor Maintenance

Blood progenitors within the lymph gland, a larval organ that supports hematopoiesis in Drosophila melanogaster, are maintained by integrating signals emanating from niche-like cells and those from differentiating blood cells. We term the signal from differentiating cells the ‘equilibrium signal’ in order to distinguish it from the ‘niche signal’. Earlier we showed that equilibrium signaling utilizes Pvr (the Drosophila PDGF/VEGF receptor), STAT92E, and adenosine deaminase-related growth factor A (ADGF-A) (<xref ref-type="bibr" rid="bib43">Mondal et al., 2011</xref>). Little is known about how this signal initiates during hematopoietic development. To identify new genes involved in lymph gland blood progenitor maintenance, particularly those involved in equilibrium signaling, we performed a genetic screen that identified bip1 (bric à brac interacting protein 1) and Nucleoporin 98 (Nup98) as additional regulators of the equilibrium signal. We show that the products of these genes along with the Bip1-interacting protein RpS8 (Ribosomal protein S8) are required for the proper expression of Pvr.

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Characterization of Cep135, a novel coiled-coil centrosomal protein involved in microtubule organization in mammalian cells

The Journal of Cell Biology ◽

10.1083/jcb.200108088 ◽

2002 ◽

Vol 156 (1) ◽

pp. 87-100 ◽

Cited By ~ 94

Author(s):

Toshiro Ohta ◽

Russell Essner ◽

Jung-Hwa Ryu ◽

Robert E. Palazzo ◽

Yumi Uetake ◽

...

Keyword(s):

Mammalian Cells ◽

Coiled Coil ◽

Amino Acid Residues ◽

Protein Overexpression ◽

Microtubule Organization ◽

Microtubule Network ◽

Centrosomal Protein ◽

Wide Range ◽

Spindle Microtubules

By using monoclonal antibodies raised against isolated clam centrosomes, we have identified a novel 135-kD centrosomal protein (Cep135), present in a wide range of organisms. Cep135 is located at the centrosome throughout the cell cycle, and localization is independent of the microtubule network. It distributes throughout the centrosomal area in association with the electron-dense material surrounding centrioles. Sequence analysis of cDNA isolated from CHO cells predicted a protein of 1,145–amino acid residues with extensive α-helical domains. Expression of a series of deletion constructs revealed the presence of three independent centrosome-targeting domains. Overexpression of Cep135 resulted in the accumulation of unique whorl-like particles in both the centrosome and the cytoplasm. Although their size, shape, and number varied according to the level of protein expression, these whorls were composed of parallel dense lines arranged in a 6-nm space. Altered levels of Cep135 by protein overexpression and/or suppression of endogenous Cep135 by RNA interference caused disorganization of interphase and mitotic spindle microtubules. Thus, Cep135 may play an important role in the centrosomal function of organizing microtubules in mammalian cells.

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maelstrom is required for an early step in the establishment of Drosophila oocyte polarity: posterior localization of grk mRNA

Development ◽

10.1242/dev.124.22.4661 ◽

1997 ◽

Vol 124 (22) ◽

pp. 4661-4671 ◽

Cited By ~ 4

Author(s):

N.J. Clegg ◽

D.M. Frost ◽

M.K. Larkin ◽

L. Subrahmanyan ◽

Z. Bryant ◽

...

Keyword(s):

Nurse Cell ◽

Egf Receptor ◽

Mrna Localization ◽

Follicle Cells ◽

Rna Transport ◽

Nurse Cells ◽

Loss Of Function ◽

Cell Fates ◽

Early Step ◽

Novel Protein

We describe a mutant, maelstrom, that disrupts a previously unobserved step in mRNA localization within the early oocyte, distinct from nurse-cell-to-oocyte RNA transport. Mutations in maelstrom disturb the localization of mRNAs for Gurken (a ligand for the Drosophila Egf receptor), Oskar and Bicoid at the posterior of the developing (stage 3–6) oocyte. maelstrom mutants display phenotypes detected in gurken loss-of-function mutants: posterior follicle cells with anterior cell fates, bicoid mRNA localization at both poles of the stage 8 oocyte and ventralization of the eggshell. These data are consistent with the suggestion that early posterior localization of gurken mRNA is essential for activation of the Egf receptor pathway in posterior follicle cells. Posterior localization of mRNA in stage 3–6 oocytes could therefore be one of the earliest known steps in the establishment of oocyte polarity. The maelstrom gene encodes a novel protein that has a punctate distribution in the cytoplasm of the nurse cells and the oocyte until the protein disappears in stage 7 of oogenesis.

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