When RNA polymerases were first studied in eukaryotes three

When RNA polymerases were first studied in eukaryotes, three peaks of polymerizing activity (1, 2 and 3 on the graph below) were obtained by fractionating cell extracts on chromatography columns (this is a common way to isolate, characterize and study cellular proteins). It was unclear whether these peaks of activity were corresponding to different RNA polymerases or just to different forms of one RNA polymerase. In an experiment, the three fractions corresponding to the enzyme activity were treated with either no, 1 (mu g/ml or 10 mu g/ml of alpha-amanitin (a highly deadly poison obtained from the mushroom Amanita phalloides). RNA polymerase activity was measured and gave the results shown in the figure below. Do these results argue for the existence of different RNA polymerases or different forms of a single RNA polymerase? Explain your answer.

Solution

It is now known that there are atleast three kinds of RNA polymerases in eukaryotes. Although preliminary studies were unable to clearly identify these subunits, but the overall effect of toxins on some of these polymerases was known. It can be seen from the image that after column chromatography, a strong and broad peak of high protein concentration was observed. Hhowever, further purification steps lead to the observation that atleast three bands were there in all the experimental group containing either none or different amount of toxin. Some points of understanding that must be noted here are:

Thus, taken together, it can be deciphered from the data that atleast three types of RNA polymerases exist in this sample. One type is that which does not get much affected due to amanitin, second one is highly prone to degradation and third one has moderate functional degradation.

It is now very well knonw that amanitin is an inhibitor of RNA transcription in vertebrates by preventing binding of RNA polymerase on the consensus sequence of DNA. Thus, the above given explanation supports the data of existence of atleast three types of RNA polymerases.