A conserved biological response to double-stranded RNA (dsRNA) mediates resistance to parasitic nucleic acids, encoded by transposable elements and viruses. Cleavage of dsRNA into small interfering RNAs (siRNAs) triggers repression of genes related by sequence to the siRNAs. This silencing response is related by mechanism to an endogenous regulatory process that represses the expression of protein-coding genes. The latter process is guided by microRNAs (miRNAs), which are produced from genes within the genome. The long-term goals of our research are to understand the molecular mechanisms of siRNA and miRNA action, and to understand what aspects of cell activities regulate silencing by small RNAs. To this end, we have developed genetic, biochemical and cell biological methods in whole Drosophila and in cells to achieve our goals. In previous work, we conducted screens for Drosophila mutations that perturb siRNA- and miRNA-mediated repression. Genes that were identified from the screens are the basis for research aims in this proposal.
Our aims are to: 1) determine how the nuclear Cdk9 kinase recruits a protein complex to co-transcriptionally cleave RNA transcripts into miRNAs, 2) determine how miRNA-protein complexes in the cytoplasm specifically repress the translation of certain mRNAs, 3) learn how these miRNA-protein complexes become associated with trafficking organelles in the cytoplasm, thereby leading to the inactivation of the miRNA-protein complexes, and 4) understand how the Dicer ribonuclease cleaves RNA from infecting viruses to generate antiviral siRNAs. The significance to health is manifold. The ability of small RNAs to regulate virus, transposon, and organismal gene expression suggests that basic understanding will reveal control mechanisms that influence many aspects of biology. It will also have an impact on viral infectious diseases and non-infectious diseases such as cancer. Moreover, small RNAs are used experimentally to probe gene function and are being commercially developed as disease therapeutics, making basic understanding of these molecules an important biomedical research goal.

Public Health Relevance

Small non-coding RNAs (siRNAs and miRNAs) are regulators of gene expression, and their dysfunction is linked to diverse kinds of human disease. Additionally, small RNAs play important defense functions during infection by numerous viruses. This project is to understand how small RNAs are made, how they control gene expression, and how they defend against virus infection.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM068743-11
Application #
8510653
Study Section
Molecular Genetics B Study Section (MGB)
Program Officer
Bender, Michael T
Project Start
2003-07-01
Project End
2015-07-31
Budget Start
2013-08-01
Budget End
2014-07-31
Support Year
11
Fiscal Year
2013
Total Cost
$333,809
Indirect Cost
$107,034
Name
Northwestern University at Chicago
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
160079455
City
Evanston
State
IL
Country
United States
Zip Code
60201
Posadas, Diana M; Carthew, Richard W (2014) MicroRNAs and their roles in developmental canalization. Curr Opin Genet Dev 27:1-6
Marques, Joao Trindade; Wang, Ji-Ping; Wang, Xiaohong et al. (2013) Functional specialization of the small interfering RNA pathway in response to virus infection. PLoS Pathog 9:e1003579
Cassidy, Justin J; Jha, Aashish R; Posadas, Diana M et al. (2013) miR-9a minimizes the phenotypic impact of genomic diversity by buffering a transcription factor. Cell 155:1556-67
Wu, Pei-Hsuan; Isaji, Mamiko; Carthew, Richard W (2013) Functionally diverse microRNA effector complexes are regulated by extracellular signaling. Mol Cell 52:113-23
Marques, Joao Trindade; Kim, Kevin; Wu, Pei-Hsuan et al. (2010) Loqs and R2D2 act sequentially in the siRNA pathway in Drosophila. Nat Struct Mol Biol 17:24-30
Carthew, Richard W; Sontheimer, Erik J (2009) Origins and Mechanisms of miRNAs and siRNAs. Cell 136:642-55
Lucchetta, Elena M; Carthew, Richard W; Ismagilov, Rustem F (2009) The endo-siRNA pathway is essential for robust development of the Drosophila embryo. PLoS One 4:e7576
Lee, Young Sik; Pressman, Sigal; Andress, Arlise P et al. (2009) Silencing by small RNAs is linked to endosomal trafficking. Nat Cell Biol 11:1150-6
Lim, Do Hwan; Kim, Jung; Kim, Sanguk et al. (2008) Functional analysis of dicer-2 missense mutations in the siRNA pathway of Drosophila. Biochem Biophys Res Commun 371:525-30
Reinke, Catherine A; Carthew, Richard W (2008) BMP signaling goes posttranscriptional in a microRNA sort of way. Dev Cell 15:174-5

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