Samples of 5 g of total RNA were reverse transcribed using M MLV reverse transcriptase and an oligo dT primer. The primers used for real time PCR were for Cyclin A2 All reactions were carried out using a 7300 Real Time PCR System and ABsolute neverless QPCR SYBR Green Mix. The amplification was carried out as follow initial enzyme activation at 94 C for 15 min, then 40 cycles of 94 C for 15 s and 60 C for 1 min. A total of 50 ng of each diluted reverse transcription product was used for real time PCR in a final volume of 25 l containing 160 nM of each specific primer and 1�� ABsolute QPCR SYBR Green Mix. The relative level of Cyclin A2 gene expression was calculated according to the comparative Inhibitors,Modulators,Libraries Ct method using the 2 CTCT formula, using the expression of Actin as an endogenous control.

Background Spinocerebellar ataxia type 8 is a dominantly inherited, slowly progressive neurodegenerative disorder caused by the expansion Inhibitors,Modulators,Libraries of CTA CTG combined repeats in the ataxin 8 opposite strand gene located on chromosome 13q21. The reported repeat lengths associated with ataxia vary dramatically, ranging from 68 to 1000 repeats. In the general popula tion more Inhibitors,Modulators,Libraries than 99% of the alleles have 16 37 CR. Nevertheless the penetrance of the SCA8 repeat expansion and ataxia is not complete, as expansions do not always segregate with ataxia in families and they are present in rare instances in normal and non ataxic diseased popula tions. The pathogenesis of SCA8 is complex. In addition to a CTG repeat expansion in the ATXN8OS gene, it also involves a CAG repeat expansion in another overlapping gene, ataxin 8.

In the CTG direction, ATXN8OS expresses non coding transcripts containing the CUG expansion which overlap with the 5 region of the Kelch like 1 transcripts, and in the CAG direction ATXN8 expresses transcripts encoding a nearly Inhibitors,Modulators,Libraries pure polyglutamine expansion protein. As a consequence, three plausible mechanisms were proposed for SCA8 RNA gain of function, partial loss of KLHL1 function and polyglutamine expansion protein in the CAG direction. In the present study, we focus on the RNA gain of function mechanism. The causative agent for myotonic dystrophy is also known to be a CTG expansion in the 3 UTR of the DMPK gene. The expanded CUG repeat in the DMPK RNA impaired nuclear cytoplasmic transport, resulting in nuclear retention and ribonuclear foci formation.

In addition, Inhibitors,Modulators,Libraries expanded CTG repeats in DM1 alter the adja cent chromatin structure and several proteins bind to CUG repeat containing RNA. Using PC12 neuro nal cells expressing the ref 1 CUG repeat bearing mRNA, cis effects through the reporter gene and neuronal death after cell differentiation in vitro were reported. Expression of a Huntingtons disease like 2 JPH3 transcript with an expanded CUG repeat also resulted in the formation of RNA foci and cell toxicity.