Application of siRNA for Targeting Neuronal Genes

Organizer: Thomas Tuschl, Rockefeller University

The goal of this group is to provide a forum for New York–area scientists engaged in research into the biology, biochemistry, and applications of RNAi silencing to meet regularly to discuss advances in this exciting new field and to promote interinstitutional and interdisciplinary dialogue. Topics of discussion include regulation of RNAi gene expression, mechanistic studies into RNAi action, regulation of gene expression by siRNAs, RNAi in health and disease, and applied uses of technologies related to RNAi.

Program

John Cryan, University College Cork, Ireland
"siRNA-Induced Widespread Gene Knockdown in the Adult Mouse Brain: Utility for Studying the Molecular Basis of Neuropsychiatric Disorders."

Sergei Musatov, Weill-Cornell Medical College
"Viral-Mediated RNA Interference in Neuroscience Research."

Richard Vallee, Columbia University
"Effects of LIS1 RNAi on Neural Stem Cell Polarization, Division, and Motility."

Stefan Schlussman, Rockefeller University
"Behavioral Effects of Local Infusion of Mu Opioid Receptor Targeted Small Interfering RNA in the Adult Mouse Brain."

Abstract

"siRNA-Induced Widespread Gene Knockdown in the Adult Mouse Brain: Utility for Studying the Molecular Basis of Neuropsychiatric Disorders"
John Cryan
We investigated whether non-viral infusion of short interfering RNA (siRNA) into the ventricular system would enable a widespread sequence-specific gene knockdown in the brain. Preliminary studies in mice, over-expressing enhanced green fluorescent protein, demonstrated an extensive, temporal and specific knockdown of this transgene in regions rostro-caudally, dorso-ventrally or medio-laterally distant to the siRNA infusion site, dorsal third ventricle. We challenged our RNAi protocol to generate a specific knockdown of two endogenous genes, coding for transporters of serotonin (SERT) and dopamine (DAT), both of which have been implicated in the etiology of neuropsychiatric disorders. Infusion of DAT or SERT-siRNAs produced behavioral effects similar to pharmacological inhibitors of these transporters. These results demonstrate the efficiency of our non-viral RNAi approach in achieving a rapid, widespread and sequence-specific gene silencing in the brain. Application of this approach may accelerate the process of target validation for neuropsychiatric disorders that involve a complex interplay of gene(s) from various brain regions.

"Viral-Mediated RNA Interference in Neuroscience Research"
Sergei Musatov
Viral vectors represent an increasingly important method for altering gene expression in vivo. In recent years, these vectors have been exploited for delivery of small hairpin RNA into the brain to achieve long-term region-specific gene silencing thus creating a powerful alternative to traditional transgenic techniques and circumventing many of their limitations. We will review recent advances in vector design for in vivo RNAi applications with focus on adeno-associated viral (AAV) vectors. To illustrate the power and challenges of this technology we will discuss recent studies aimed at dissecting the role of estrogen receptor alpha in regulating specific behaviors.

"Effects of LIS1 RNAi on Neural Stem Cell Polarization, Division, and Motility"
Richard Vallee, Columbia University
Mutations in the human LIS1 gene cause classical lissencephaly ( smooth brain), which has been proposed to result from a neuronal migration. disorder. LIS1 functions in the cytoplasmic dynein pathway, but its cellular role is only partially understood. To explore the detailed mechanisms underlying this disease, we conducted the first in situ live-cell imaging analysis of LIS1-deficient neural progenitors throughout the entire radial migration pathway (Tsai, J-W, Chen