Autoregulation of Fox protein expression to produce dominant negative splicing factors

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FIGURE 5.
FIGURE 5.

Fox-2ΔE6 inhibits Fox-dependent activation of splicing. (A) HEK293 cells were transfected with the minigene DUP-E33CACNA1Cwt, containing a Fox-dependent alternative middle exon. A constant amount of FLAG-Fox-2 was cotransfected with increasing amounts of FLAG-Fox-2ΔE6. (Lane −) A control transfection with the minigene only was performed in parallel. (Top) The inclusion of the alternative exon was determined by RT-PCR. The percent exon inclusion is given below. (Middle) The amounts of FLAG-Fox-2 and FLAG-Fox-2ΔE6 were measured by anti-FLAG western. (Bottom) Anti-U1 70K Western served as a loading control. The relative FLAG-Fox-2 levels, shown below each lane are normalized to the U1-70K content. The molar ratios of Fox-2 to Fox-2ΔE6 are indicated. The effect of Fox-2ΔE6 on Fox-1-dependent and Fox-3-dependent splicing of the DUP-E33CACNA1Cwt minigene was studied in a similar way and is shown in B and C, respectively.

This Article

  1. Published in Advance December 30, 2009, doi: 10.1261/rna.1838210 RNA 16: 405-416

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