We are studying how the cyclic AMP receptor protein (CRP) activates transcription in Escherichia coli. (i) We have found that CRP activation of lac transcription not only requires protein-protein interaction with RNA polymerase, but also transmission of some signal through the DNA to the promoter. We have shown that for this signal to be transmitted, DNA must be intact with proper Watson-Crick base-pairing. (ii) Using both genetic and biochemical approaches, we have also found that CRP needs some accessory factors for full activation capability. One of these factors is adenylate cyclase. We have shown that when cAMP is replaced by adenylate cyclase and ATP, CRP activates transcription of the lac promoter 25-fold compared to 5-fold stimulation seen in the presence of cAMP. We have proposed that adenylate cyclase interacts with CRP during which time cAMP is translocated to CRP. (iii) The pts operon encodes proteins necessary for uptake of many sugars and for activation of adenylate cyclase. We have found that the genetic regulation of pts is quite unusual: There are multiple promoters each with multiple initiation sites that are regulated by the presence of cAMP and CRP, the degree of supercoiling of the DNA, and by alternate sigma factors.
