You are interested in identifying important transcription fa

You are interested in identifying important transcription factors for the beta-globin gene and therefore perform two experiments: 1) A linker scanning assay in which you use Northern blotting to monitor the effect on transcription of inserting 10 bp mutations centered around positions -55 to +5 of the promoter as indicated. (\'WT\' = wild-type). 2) A DNase footprinting assay for the promoter region from position -60 to +10, ^32P-labeled at the -60 position, incubated with four DNA binding proteins: a, b, c and d. (\'-\': no protein) 4b) Based on assays 1 and 2 on the right, for each of proteins a-d indicate which position in the promoter is the center of their binding site, and whether they are activators or repressors of transcription. (If a protein does not bind and/or affect transcription, write \'None\' in the respective column).

Solution

From the DNase footprinting assay it is clear that transcription factor \'a\' binds around -35bp (-30 to -40bp), as we dont find any DNA bands at that region. Binding of the transcription factor, prevents the DNase to attack and cleave that region of DNA. Similarly, \'c\' binds around -45bp and \'d\' binds around -25bp and \'b\' doesnot bind to the DNA.

Northern blotting is used for detecting the RNA. From the northern blot, it is clear that binding of a transcription factor around -35bp region enhances the mRNA production, which indicates that this transcription factor is an activator. Therefore, we could say that \'a\' is an activator. On the other hand, binding of the transcription factor around -25bp reduces the producion of mRNA and thereby reduces the expression. And that transcription factor is a repressor. i.e. protein \'d\' is a repressor.

Binding of proein \'c\' around -45 bp region doesnot alter the expression level and protein \'b\' doesnot bind to the given DNA sequence.