Our long-term goal is to understand the molecular and biochemical mechanisms of RNA interference (RNAi), a conserved post-transcriptional gene-silencing mechanism mediated by microRNA (miRNA) and small interfering RNA (siRNA). These tiny regulatory RNAs play critical roles in many fundamental biological processes. In particular, misregulation of miRNAs has recently been linked to human diseases, such as cancer. Moreover, RNAi has been widely used as a powerful gene-silencing tool to performfunctional genomic studies in multiple model systems. The specificity and potency of RNAi highlight its potential for developing novel and efficient therapeutic avenues to treat human diseases. We have previously identified two Dicer complexes, DCR-1/R3D1and DCR-2/R2D2,as the respectivemiRNA- and siRNA-generating enzymes in Drosophila. In addition, the DCR-2/R2D2 complex binds nascent siRNA and facilitates its incorporation into the siRNA-induced silencing complex (siRISC). Based on these studies, the goal of this proposal is to address several outstanding questions in the Drosophila RNAi pathway with an emphasis on the miRNA pathway. We will investigate the inherent specificity and mechanism of miRNA biogenesis (Aim 1). We will test our hypothesis that the DCR-1/R3D1 complex senses asymmetry of miRNA and facilitates miRNA loading onto its effector complex, miRISC (Aim 2). Finally, we will develop a reconstitution assay to explore the mechanism of miRISC assembly and to identify and characterize new components of miRISC assembly (Aim 3). These studies will significantly advance our understanding of the Drosophila RNAi pathway. The origins of human diseases, such as cancer, can be generally attributed to loss-of-function of important genes (tumor suppressor genes) and/or gain-of-function of pathological genes (oncogenes). For unknown reasons, RNAi in human cells is less efficient than RNAi in Drosophila cells. Thus, comparison of the Drosophila and human systems will allow us to identify the underlying differences and, hopefully, to optimize RNAi in human cells. This could lead to the development of novel and efficient small RNAs-based gene-silencing methods to cure human diseases by specifically shutting down pathological genes.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
3R01GM078163-04S1
Application #
7887184
Study Section
Molecular Genetics A Study Section (MGA)
Program Officer
Bender, Michael T
Project Start
2009-08-03
Project End
2010-07-31
Budget Start
2009-08-03
Budget End
2010-07-31
Support Year
4
Fiscal Year
2009
Total Cost
$80,000
Indirect Cost
Name
University of Texas Sw Medical Center Dallas
Department
Biochemistry
Type
Schools of Medicine
DUNS #
800771545
City
Dallas
State
TX
Country
United States
Zip Code
75390
Norton, John T; Huang, Sui (2013) The perinucleolar compartment: RNA metabolism and cancer. Cancer Treat Res 158:139-52
Welker, Noah C; Maity, Tuhin S; Ye, Xuecheng et al. (2011) Dicer's helicase domain discriminates dsRNA termini to promote an altered reaction mode. Mol Cell 41:589-99
Liu, Ying; Tan, Huiling; Tian, Hui et al. (2011) Autoantigen La promotes efficient RNAi, antiviral response, and transposon silencing by facilitating multiple-turnover RISC catalysis. Mol Cell 44:502-8
Yoda, Mayuko; Kawamata, Tomoko; Paroo, Zain et al. (2010) ATP-dependent human RISC assembly pathways. Nat Struct Mol Biol 17:17-23
Paroo, Zain; Ye, Xuecheng; Chen, She et al. (2009) Phosphorylation of the human microRNA-generating complex mediates MAPK/Erk signaling. Cell 139:112-22
Liu, Ying; Ye, Xuecheng; Jiang, Feng et al. (2009) C3PO, an endoribonuclease that promotes RNAi by facilitating RISC activation. Science 325:750-3
Ye, Xuecheng; Liu, Qinghua (2008) Expression, purification, and analysis of recombinant Drosophila Dicer-1 and Dicer-2 enzymes. Methods Mol Biol 442:11-27
Kalidas, Savitha; Sanders, Charcacia; Ye, Xuecheng et al. (2008) Drosophila R2D2 mediates follicle formation in somatic tissues through interactions with Dicer-1. Mech Dev 125:475-85
Liu, Xiang; Park, Joseph K; Jiang, Feng et al. (2007) Dicer-1, but not Loquacious, is critical for assembly of miRNA-induced silencing complexes. RNA 13:2324-9
Ye, Xuecheng; Paroo, Zain; Liu, Qinghua (2007) Functional anatomy of the Drosophila microRNA-generating enzyme. J Biol Chem 282:28373-8

Showing the most recent 10 out of 12 publications