Cytoplasmic RNA localization is a widespread mechanism for generating both embryonic and somatic cell polarity. Among vertebrates, Vg1 mRNA is a prominent example of a localized mRNA that plays a role in embryonic patterning. Vg1 mRNA encodes a peptide growth factor, and is localized during oogenesis to the vegetal cytoplasm of Xenopus oocytes. Restricted expression of Vg1 protein in the vegetal hemisphere of the egg is critical for correct patterning of the embryo, making localization of Vg1 mRNA an important model for understanding how maternal molecules are localized to influence pattern and polarity. The goal of this research project is to investigate how RNA molecules can be targeted to specific regions of the cell cytoplasm to generate cell polarity through spatially restricted protein expression. The foundation for this investigation has been laid by our progress studying the molecular interactions that govern this process. Our experiments have defined critical RNA-protein and protein-protein interactions leading to assembly of a transport-competent RNP complex and have uncovered distinct steps in the localization pathway. Our current proposal seeks to extend these findings by studying the dynamic transitions between these steps in the localization pathway. We will use a combination of in vivo and in vitro approaches to determine the molecular mechanisms responsible for localized expression of Vg1 mRNA.
Three specific aims are proposed:
In Aim I, we will characterize the dynamics and transport of the cytoplasmic Vg1 RNP complex, in Aim II, we will analyze the assembly and activity of RNP transport granules and in Aim III we will investigate the mechanisms that couple translational repression and RNA localization The proposed research is designed to reveal the mechanisms by which mRNA molecules are transported within cells to generate spatially restricted protein expression, and will provide insight into how developmental signals are spatially distributed in the vertebrate embryo. This work will impact issues related to human health such as birth defects, as well as neurological diseases that have been linked to defective RNA transport and localized protein synthesis.

Public Health Relevance

The goal of this research project is to elucidate the molecular mechanisms responsible for RNA localization and localized protein synthesis. These processes are critical for both embryonic development and neurological function. In particular, certain human diseases, including fragile X mental retardation syndrome and spinal muscular atrophy, have been linked to defects in RNA localization and localized protein synthesis.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM071049-17
Application #
8197643
Study Section
Development - 2 Study Section (DEV2)
Program Officer
Hoodbhoy, Tanya
Project Start
1993-09-01
Project End
2014-11-30
Budget Start
2011-12-01
Budget End
2014-11-30
Support Year
17
Fiscal Year
2012
Total Cost
$356,644
Indirect Cost
$131,221
Name
Brown University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
001785542
City
Providence
State
RI
Country
United States
Zip Code
02912
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Pratt, Catherine A; Mowry, Kimberly L (2013) Taking a cellular road-trip: mRNA transport and anchoring. Curr Opin Cell Biol 25:99-106
Gagnon, James A; Mowry, Kimberly L (2011) Visualization of mRNA localization in Xenopus oocytes. Methods Mol Biol 714:71-82
Gagnon, James A; Mowry, Kimberly L (2011) Molecular motors: directing traffic during RNA localization. Crit Rev Biochem Mol Biol 46:229-39
Gagnon, James A; Mowry, Kimberly L (2010) Visualizing RNA localization in Xenopus oocytes. J Vis Exp :
Pratt, Catherine A; Mowry, Kimberly L (2010) Preparation of a highly active cell-free translation system from immature Xenopus laevis oocytes. Methods 51:101-5
Gagnon, James A; Mowry, Kimberly L (2009) VISIONS: the art of science. Mol Reprod Dev 76:1115
Messitt, Timothy J; Gagnon, James A; Kreiling, Jill A et al. (2008) Multiple kinesin motors coordinate cytoplasmic RNA transport on a subpopulation of microtubules in Xenopus oocytes. Dev Cell 15:426-36
Lewis, Raymond A; Gagnon, James A; Mowry, Kimberly L (2008) PTB/hnRNP I is required for RNP remodeling during RNA localization in Xenopus oocytes. Mol Cell Biol 28:678-86