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C2 P1. PTB. p50. BAG-1. p46. p36. Actin. 1.2. 1. 0.8. p1. 0.6. Relative luciferase activity. c2. 0.4. 0.2. 0. pRF. pRBagF. Supplementary data 1. HeLa cells were transfected with siRNAs directed against PTB P1

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C2 P1

PTB

p50

BAG-1

p46

p36

Actin

1.2

1

0.8

p1

0.6

Relative luciferase activity

c2

0.4

0.2

0

pRF

pRBagF

Supplementary data 1.

HeLa cells were transfected with siRNAs directed against PTB P1

(AAC UUC CAU CAU UCC AGA GAA) or a control sequence and C2 (AAG GUC CGG CUC CCC CAA AUG)

as described previously (Mitchell et al 2005). Cells were then transfected with the di-cistronic plasmid pRBagF (as

described; Coldwell et al 2000) or the control plasmid pRF. Lysates were prepared from transfected cells

and luciferase activities were measured and calculated as previously described (Mitchell et al., 2003) using

a Dual-Luciferase reporter assay system (Promega). Western analysis was performed on these samples to

detect expression of PTB, and BAG-1 whilst actin was used as a loading control. It can be seen that when PTB levels are reduced there is a specific reduction in the level of the p36 isoform of BAG-1 and a corresponding reduction in BAG-1 IRES activity.


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