Chemical Modifications of siRNA Bases

Selective nuclease digestion of messenger RNAs inside living cells via the short interfering RNA (siRNA)-triggered RNA interference pathway has become a mainstay in molecular biology to study gene function, and it holds promise for the development of new therapeutics.  However, gaps in our understanding of the basic RNA interference mechanism and ability of siRNAs to interact with intracellular RNA-binding proteins, particularly those involved in the innate immune response, limit the application of this promising new technology.  Our laboratory is developing new synthetic approaches to modified RNA oligonucleotides that carry out gene silencing with reduced undesirable off-target effects.  Specifically, we are engaged in the study of alkylated RNA bases that change the shape of the minor or major groove of double-stranded RNA while maintaining Watson-Crick-like base pairing.  These siRNA duplexes are being used to determine the effect of steric blocks in gene silencing.  The modifications are also expected to block sequence-dependent off-target ettects mediated by Toll-like receptors andsequence-independent effects from dsRBM proteins.

Secondly, we are developing modified bases capable to switching their steric blockades from the minor groove to the major groove (see figure).  These modifications will be placed in the antisense strand using a Watson-Crick-paired sense strand for delivery; the antisense strand will be designed to target a complementary mRNA sequence via Hoogsteen base pairing in which a conformational change hides the steric blockade in the major groove.
This project is being conducted in collaboration with Professor Peter Beal (UC, Davis) and is supported by the NIH, R01 GM080784.

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