RNA recognition by p53. (A) The yeast three-hybrid system. Transcription of LacZ and HIS3 reporter genes integrated at independent sites in the yeast genome depends on a trimolecular interaction between hybrid 1 (LexA/MS2 coat protein fusion); hybrid 2, an RNA comprised of one or more MS2 recognition sequences (gray) and a test RNA sequence; and hybrid 3 (GAL4AD/p53 protein fusion). (B) Predicted secondary structures of hybrid RNAs tested in a preliminary yeast three-hybrid system assay. The RNA designated –/(MS2)2 encodes two copies of the MS2 recognition sequence (gray), as well as RNase P RPR1 leader and terminator sequences (SenGupta et al. 1996). In addition to the sequences present in the –/(MS2)2 RNA, α-p50/(MS2)2 encodes an in vitro-selected RNA aptamer targeted to transcription factor NF-κB (Cassiday and Maher 2001), and IRE/(MS2)2 encodes the iron response element RNA (Sen-Gupta et al. 1996). (C) Colony filter-lift assay for LacZ reporter gene expression in the yeast three-hybrid system. Two independent transformants of each yeast strain are shown.
