ss-siRNAs allele selectively inhibit ataxin-3 expression: multiple mechanisms for an alternative gene silencing strategy.
Single-stranded silencing RNAs (ss-siRNAs) provide an alternative approach to gene silencing. ss-siRNAs combine the simplicity and favorable biodistribution of antisense oligonucleotides with robust silencing through RNA interference (RNAi). Previous studies reported potent and allele-selective inhibition of human huntingtin expression by ss-siRNAs that target the expanded CAG repeats within the mutant allele. Mutant ataxin-3, the genetic cause of Machado-Joseph Disease, also contains an expanded CAG repeat. We demonstrate here that ss-siRNAs are allele-selective inhibitors of ataxin-3 expression and then redesign ss-siRNAs to optimize their selectivity. We find that both RNAi-related and non-RNAi-related mechanisms affect gene expression by either blocking translation or affecting alternative splicing. These results have four broad implications: (i) ss-siRNAs will not always behave similarly to analogous RNA duplexes; (ii) the sequences surrounding CAG repeats affect allele-selectivity of anti-CAG oligonucleotides; (iii) ss-siRNAs can function through multiple mechanisms and; and (iv) it is possible to use chemical modification to optimize ss-siRNA properties and improve their potential for drug discovery.
Departments of Pharmacology and Biochemistry, UT Southwestern Medical Center at Dallas, Dallas, TX 75390, USA and Department of Medicinal Chemistry and Core Antisense Research, ISIS Pharmaceuticals, Carlsbad, CA 92010, USA.
Jing
J
Liu
Dongbo
D
Yu
Yuichiro
Y
Aiba
Hannah
H
Pendergraff
Eric E
EE
Swayze
Walt F
WF
Lima
Jiaxin
J
Hu
Thazha P
TP
Prakash
David R
DR
Corey
D000483:Alleles; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000067699:Ataxin-3; D002460:Cell Line; D006801:Humans; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics*; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000737:chemistry*; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics*; D018911:Trinucleotide Repeats
D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D012097:Repressor Proteins; C092341:ATXN3 protein, human; D000067699:Ataxin-3