Abstract

application): The PI proposes to study all posttranscriptional steps involved in biogenesis of the U3 small nucleolar ribonucleoprotein particle (U3 snoRNP). These steps include binding of proteins to the precursor and mature forms of U3 snoRNA; hypermethylation of the 5' cap; nuclear retention of the RNA, RNP intermediates, and the mature snoRNP; and finally, relocalization of the U3 snoRNP from the nucleoplasm to the nucleolus. The experimental system of choice will be the Xenopus oocyte for all the usual reasons. In particular, microinjected U3 snoRNAs are reconstituted into U3 snoRNPs, and the subcellular localization of the resulting RNPs (cytoplasm, nucleoplasm, nucleoli) are easily determined both by biochemical fractionation and by traditional light microscopy. The Xenopus assay system will be used to determine the protein composition of reconstituted U3 snoRNPs, and the role of various U3 snoRNA sequences in protein binding, snoRNP reconstitution, and nucleolar localization. U3 snoRNP proteins will be identified by analytical assays (crosslinking, affinity selection) but cloned by two hybrid screens (for those that might interact with fibrillarin) and three hybrid screens (for those that might interact directly with U3 snoRNA).

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM054682-04
Application #
6181108
Study Section
Molecular Biology Study Section (MBY)
Program Officer
Rhoades, Marcus M
Project Start
1997-08-01
Project End
2002-07-31
Budget Start
2000-08-01
Budget End
2001-07-31
Support Year
4
Fiscal Year
2000
Total Cost
$190,692
Indirect Cost

  Institution

Name
University of Georgia
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
City
Athens
State
GA
Country
United States
Zip Code
30602

  Publications

Elmore, Joshua R; Sheppard, Nolan F; Ramia, Nancy et al. (2016) Bipartite recognition of target RNAs activates DNA cleavage by the Type III-B CRISPR-Cas system. Genes Dev 30:447-59
Sheppard, Nolan F; Glover 3rd, Claiborne V C; Terns, Rebecca M et al. (2016) The CRISPR-associated Csx1 protein of Pyrococcus furiosus is an adenosine-specific endoribonuclease. RNA 22:216-24
Makarova, Kira S; Wolf, Yuri I; Alkhnbashi, Omer S et al. (2015) An updated evolutionary classification of CRISPR-Cas systems. Nat Rev Microbiol 13:722-36
Swarts, Daan C; Hegge, Jorrit W; Hinojo, Ismael et al. (2015) Argonaute of the archaeon Pyrococcus furiosus is a DNA-guided nuclease that targets cognate DNA. Nucleic Acids Res 43:5120-9
Elmore, Joshua; Deighan, Trace; Westpheling, Jan et al. (2015) DNA targeting by the type I-G and type I-A CRISPR-Cas systems of Pyrococcus furiosus. Nucleic Acids Res 43:10353-63
Majumdar, Sonali; Zhao, Peng; Pfister, Neil T et al. (2015) Three CRISPR-Cas immune effector complexes coexist in Pyrococcus furiosus. RNA 21:1147-58
Hale, Caryn R; Cocozaki, Alexis; Li, Hong et al. (2014) Target RNA capture and cleavage by the Cmr type III-B CRISPR-Cas effector complex. Genes Dev 28:2432-43
Terns, Rebecca M; Terns, Michael P (2014) CRISPR-based technologies: prokaryotic defense weapons repurposed. Trends Genet 30:111-8
Ramia, Nancy F; Spilman, Michael; Tang, Li et al. (2014) Essential structural and functional roles of the Cmr4 subunit in RNA cleavage by the Cmr CRISPR-Cas complex. Cell Rep 9:1610-1617
Shao, Yaming; Cocozaki, Alexis I; Ramia, Nancy F et al. (2013) Structure of the Cmr2-Cmr3 subcomplex of the Cmr RNA silencing complex. Structure 21:376-84

Showing the most recent 10 out of 41 publications