description) This application proposes to test the hypothesis that fragile-X mental retardation results from a failure of FMRP to bind sequence-specific RNA targets, and to identify those target RNAs. Three aspects of this issue will be definitively addressed. First, FMRP sequence-specific RNA targets will be identified using in vitro RNA selection methods, specifically delineating RNAs that bind to full-length FMRP and to the FMRP KH2 RNA binding domain, in which an I to N mutation is implicated in fragile-X mental retardation. In addition, mutagenesis studies will yield insight into the nucleic acid and amino acid residues that are critical for RNA-protein recognition. Second, the investigators will utilize this information to identify candidate in vivo FMRP RNA targets, using several redundant strategies. Third, they will test the metabolism of these candidate RNAs in vivo in genetically defined FMR null mice. The investigators will utilize FMR null mice as well BAC transgenic technology to rescue the FMR-1 null mice with FMR-1 genes harboring specific mutations in KH2 necessary for sequence-specific RNA binding. Examination of phenotype of mice harboring KH2 point-mutants will allow the investigators to assign aspects of neuronal dysfunction that result from failures in sequence-specific RNA binding. Moreover, comparison of the metabolism of candidate RNAs in mutant with wild-type mice will allow the investigators to conclude that FMRP acts on specific RNA targets in neurons.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD040647-04
Application #
6743944
Study Section
Special Emphasis Panel (ZHD1-MRG-C (15))
Program Officer
Vitkovic, Ljubisa
Project Start
2001-04-15
Project End
2006-03-31
Budget Start
2004-04-01
Budget End
2006-03-31
Support Year
4
Fiscal Year
2004
Total Cost
$297,250
Indirect Cost
Name
Rockefeller University
Department
Internal Medicine/Medicine
Type
Other Domestic Higher Education
DUNS #
071037113
City
New York
State
NY
Country
United States
Zip Code
10065
Moore, Michael J; Zhang, Chaolin; Gantman, Emily Conn et al. (2016) Erratum: Mapping Argonaute and conventional RNA-binding protein interactions with RNA at single-nucleotide resolution using HITS-CLIP and CIMS analysis. Nat Protoc 11:616
Vasilyev, Nikita; Polonskaia, Anna; Darnell, Jennifer C et al. (2015) Crystal structure reveals specific recognition of a G-quadruplex RNA by a ?-turn in the RGG motif of FMRP. Proc Natl Acad Sci U S A 112:E5391-400
Berry-Kravis, Elizabeth; Levin, Rebecca; Shah, Haroon et al. (2015) Cholesterol levels in fragile X syndrome. Am J Med Genet A 167A:379-84
Moore, Michael J; Zhang, Chaolin; Gantman, Emily Conn et al. (2014) Mapping Argonaute and conventional RNA-binding protein interactions with RNA at single-nucleotide resolution using HITS-CLIP and CIMS analysis. Nat Protoc 9:263-93
Darnell, Jennifer C; Klann, Eric (2013) The translation of translational control by FMRP: therapeutic targets for FXS. Nat Neurosci 16:1530-6
Teplova, Marianna; Malinina, Lucy; Darnell, Jennifer C et al. (2011) Protein-RNA and protein-protein recognition by dual KH1/2 domains of the neuronal splicing factor Nova-1. Structure 19:930-44
Darnell, J C (2011) Defects in translational regulation contributing to human cognitive and behavioral disease. Curr Opin Genet Dev 21:465-73
Darnell, Jennifer C; Van Driesche, Sarah J; Zhang, Chaolin et al. (2011) FMRP stalls ribosomal translocation on mRNAs linked to synaptic function and autism. Cell 146:247-61
Phan, Anh Tuan; Kuryavyi, Vitaly; Darnell, Jennifer C et al. (2011) Structure-function studies of FMRP RGG peptide recognition of an RNA duplex-quadruplex junction. Nat Struct Mol Biol 18:796-804
Darnell, Jennifer C; Fraser, Claire E; Mostovetsky, Olga et al. (2009) Discrimination of common and unique RNA-binding activities among Fragile X mental retardation protein paralogs. Hum Mol Genet 18:3164-77

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