DISTANCE CONSTRAINTS AND STEREOSPECIFIC ALIGNMENT REQUIREMENTS CHARACTERISTIC OF P53 DNA-BINDING CONSENSUS SEQUENCE HOMOLOGIES

We present evidence in favor of the position that some mutant p53 prot eins retain the ability to trans-activate downstream genes through p53 DNA-binding consensus sequence (CS) homologies. We tested one cell li ne possessing high levels of mutant p53 and found that this mutant p53 is highly active in trans-activating one CS homology, moderately acti ve in trans-activating a second sequence and inactive in modulating a third sequence. We tested a second cell line, also possessing high lev els of mutant p53 and found the same pattern of activation. In additio n we find that inter-motif distance [represented by N in RRRCWWGYYY(N) RRRCWWGYYY] is very important in determining the relative binding affi nity of a given CS homology for wild-type or mutant p53. Our studies s uggest that stereospecific alignment of the DNA-binding motifs within the CS may favor binding of wildtype p53 while misalignment may favor binding of mutant p53. Furthermore, we find that the maximum distances at which p53 DNA-binding CS homologies are functionally active vary f or different sequences. Introduction of as few as 200 bp between one C S homology and the downstream TATA box can eliminate a 45-fold p53-med iated trans-activation. We present evidence that the composition of th e DNA which flanks a p53 DNA-binding consensus sequence may also modul ate trans-activation.