A monoclonal antibody directed against a small subunit of RNA polymerase B blocks the initiation step

Abstract

Monoclonal antibodies directed against RNA polymerase B of the fungus Podospora were probed for their inhibitory effect on enzyme activity in vitro. From nine antibodies directed against various subunits of the enzyme only two of them totally inhibit RNA polymerase B activity in a non-selective transcription system. Enzymes A and C are not inhibited at all. The two antibodies recognize the same small subunit (B11) of Podospora enzyme B. One of these antibodies has been used as a specific inhibitor to study the function of the B11 subunit. Enzyme activity is partially protected from the inhibition when the enzyme is previously incubated with the template. The antibody does not interfere with enzyme-DNA complex formation but blocks the initiation process in a transcription system using a dinucleotide as initiator. Elongation of RNA chains does not appear to be affected. Also the antibody cross-reacts with yeast and calf thymus RNA polymerase B and has some, but less, effect on initiation by these enzymes. It is suggested the antigenic site corresponding to this antibody may overlap the catalytic initiation site on the native form of RNA polymerase B.