Book of Abstracts: Albany 2009
June 16-20 2009
© Adenine Press (2008)
CAG repeat hairpins as potential triggers of RNA-mediated pathogenesis and therapeutic targets in polyglutamine diseases
The tandem repeats of various trinucleotide motifs are abundant in human genes and many of the repeats are retained in mature transcripts. The repeats are present in the translated and untranslated regions of mRNAs, some of them are polymorphic in length and may play regulatory roles in gene expression. The CAG repeats which are the focus of our present research belong to the most frequent triplet repeats and nearly 200 mRNAs contain their tracts composed of at least six repeated units. These repeats are often translated to polyglutamine tracts in proteins and their expansion beyond the normal range in some genes result in incurable neurodegenerative disorders known as ?polyglutamine diseases?. Examples of such disorders are Huntington?s disease (HD) and a number of spinocerebellar ataxias (SCA).
Considerable information has been gathered regarding the length polymorphism and structures formed by the CAG repeat tracts in human transcripts. The CAG repeat as compared to other CNG repeats forms the least stable hairpins due to the A-A mismatches separating every two G-C and C-G base pairs in the CAG repeat hairpin stem. These hairpins are further destabilized by the repeat interruptions present in many normal alleles of the SCA1, SCA2 and SCA17 related transcripts. Other CAG repeat containing transcripts such as FOXP2 also contain repeat interruptions. The formation of split hairpins in these mRNAs which are translated to proteins having long polyglutamine tracts (>40Q) but not giving rise to any diseases may be considered a good argument for the contribution of CAG repeat hairpin toxicity in polyglutamine diseases. We provide experimental data showing that the impaired alternative splicing of some transcripts may be involved in such toxicity.
We also studied the process of RNA interference between the CAG repeats present in the endogenous transcripts of polyglutamine disease genes and exogenous reagents containing complementary CUG repeats. These reagents were either double-stranded or single-stranded siRNAs or vector-based shRNAs releasing reagents composed of repeats. We analyzed the silencing effects from various perspectives: that of silencing reagents and their targets as well as from the perspectives of the silencing reactions and their products. We intended to learn more about the efficiency and specificity of these processes and investigate the possible interplay between RNA interference and antisense mechanisms. We also wanted to explore the potential of the repeat targeting strategies for the therapy of polyglutamine diseases. The results of these studies will be presented and discussed.
Institute of Bioorganic Chemistry