Despite its novelty in biological research, the involvement of RNA interference (RNAi) as a means of gene regulation has been established in numerous natural and biotechnology systems.(3,5-6,10,11,35) When perturbed in a cellular context, RNAi has been linked to diseases such as cancer;however, due to its general mode of action also shows promise as an adaptable therapeutic tool for a number of genetic diseases. While substantial progress has been made in this relatively young field, much remains unknown about the enzymatic mechanisms that underlie RNAi. This is especially evident in the case of Piwi RNA action, a more recently discovered class of RNAi which specifically regulates gene expression through the use of small RNAs ~24-31 nucleotides in length.(9-11) Bioinformatics analysis of the Piwi-related proteins has suggested key similarities and differences among individual members of this group including their mechanism of action. Structural information for Argonautes is rather limited, however, greatly hindering dissection of their modulated functions. Therefore, this proposal aims to determine the structural basis for RNA interference in the Piwi clade of Argonaute proteins. Given the strong structural component to the proposed research, the primary methods to be employed will be recombinant protein expression, purification, crystallization, and structure determination by X-ray diffraction. Once a structure solution has been determined, thorough biophysical and biochemical analysis of the protein will follow based on observations made from the structure. Overall, this research plan will elucidate many details of RNAi action and provide insight into RNA interference mechanisms. Given the links between RNAi, diseases such as cancer, and therapeutics, understanding the specifics of RNAi biology will enhance our understanding of the molecular basis of disease and assist in the development of more effective therapies.

Public Health Relevance

This work will provide insight into the mechanism of RNA interference (RNAi) through biochemical and structural studies. Given the links between RNAi, diseases such as cancer, and therapeutics, understanding the specifics of RNAi biology will enhance our understanding of the molecular basis of disease and assist in the development of more effective therapies.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32GM097888-02
Application #
8335565
Study Section
Special Emphasis Panel (ZRG1-F04B-D (20))
Program Officer
Flicker, Paula F
Project Start
2011-09-01
Project End
2013-08-31
Budget Start
2012-09-01
Budget End
2013-08-31
Support Year
2
Fiscal Year
2012
Total Cost
$52,190
Indirect Cost
Name
Cold Spring Harbor Laboratory
Department
Type
DUNS #
065968786
City
Cold Spring Harbor
State
NY
Country
United States
Zip Code
11724