The long-term objective of this research is to elucidate the mRNA translational regulatory mechanisms that control cellular morphogenesis in cleavage stage Drosophila embryos. Embryogenesis in all animals begins with a cleavage stage when cells proliferate through division without growing and undergo a maternal to zygotic transition (MZT) in gene expression. Despite the fundamental importance of this evolutionarily conserved phase of development, the mRNA translational regulatory mechanisms that function during the MZT to specifically control cellular morphogenesis are poorly understood. We have found that Drosophila Fragile X mental retardation protein (dFMRP), a translational regulator, is required for normal cell formation in cleavage stage embryos. In humans, low FMRP activity causes Fragile X syndrome (FXS), the most common form of heritable mental retardation and autism. The precise mechanism of FMRP translational regulation is uncertain. We have shown that the expression of trailer hitch (tral) mRNA, which encodes a second translational regulator, is a direct target of dFMRP-dependent regulation and that dFMRP and TRAL function during the MZT within dynamic cytoplasmic ribonucleoprotein (RNP) bodies (MZT bodies). Elucidating the functional properties of these MZT bodies and identifying the mRNAs that they regulate will significantly enhance our understanding of the molecular mechanisms controlling cellular morphogenesis in cleavage stage embryos. Using Drosophila as a model, we will conduct a comprehensive analysis of the dFMRP-dependent translational regulatory mechanisms that control cellular morphogenesis. We will use proteomic screens to identify proteins misexpressed in dfmr1 and tral mutant embryos and RNA-binding assays to determine which of the corresponding mRNAs are potentially direct targets of dFMRP in vivo. Genetic analysis will determine which identified proteins are essential for cellular morphogenesis. With these findings we will establish a translational regulatory pathway that controls cellular morphogenesis. We will also determine the function of the MZT bodies. The protein components of MZT bodies will be identified through immunofluorescence localization of known RNP body (e.g., stress granule) markers. To determine the mechanism by which dFMRP regulates expression of specific mRNAs within MZT bodies we will use a combination of in vitro translation and polyribosome-association assays, and high resolution fluorescence in situ hybridization to localize mRNAs directly bound by dFMRP. These mechanistic studies will be complemented by the biochemical and genetic characterization of a newly identified dFMRP-associated protein that is implicated in translational initiation and required for cell division cycle control in cleavage stage embryos. Our study will provide valuable insights into the molecular mechanisms that control early animal development and the etiology of FXS that could point to new therapies for FXS.

Public Health Relevance

Low FMR protein activity causes Fragile X syndrome (FXS), the most common form of heritable mental retardation and autism in humans. We have found that the Drosophila (fruit fly) counterpart of this protein is required for an early stage of embryonic development that is evolutionarily conserved in all animals. Our experimental plan to elucidate the mechanism of Drosophila FMR protein function should provide meaningful insights into the causes of FXS that could lead to new treatments for FXS in children, and advance our basic understanding of the molecular mechanisms that control early animal development.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM087562-01
Application #
7636155
Study Section
Development - 1 Study Section (DEV1)
Program Officer
Haynes, Susan R
Project Start
2009-04-01
Project End
2013-03-31
Budget Start
2009-04-01
Budget End
2010-03-31
Support Year
1
Fiscal Year
2009
Total Cost
$293,082
Indirect Cost
Name
University of Texas Austin
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
170230239
City
Austin
State
TX
Country
United States
Zip Code
78712
Reich, John; Papoulas, Ophelia (2012) Caprin controls follicle stem cell fate in the Drosophila ovary. PLoS One 7:e35365
Monzo, Kate; Dowd, Susan R; Minden, Jonathan S et al. (2010) Proteomic analysis reveals CCT is a target of Fragile X mental retardation protein regulation in Drosophila. Dev Biol 340:408-18