C. Giardina et Jt. Lis, POLYMERASE PROCESSIVITY AND TERMINATION ON DROSOPHILA HEAT-SHOCK GENES, The Journal of biological chemistry, 268(32), 1993, pp. 23806-23811
5,6-Dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) limits RNA poly
merase II transcription to a gene's 5'-end. Transcription of the unind
uced Drosophila hsp70 gene is likewise restricted to the 5'-end, where
the polymerase resides in a paused state. Furthermore, paused elongat
ion complexes formed on the uninduced hsp70 gene and in DRB-inhibited
reactions can both be restarted by Sarkosyl or high salt. These simila
rities prompted us to explore whether these complexes were generated b
y a block at the same polymerase modification step. In vivo UV cross-l
inking and KMnO4 hyperreactive site mapping show that while the natura
lly paused polymerase is restricted to the first approximately 42 base
pairs of hsp70, DRB treatment of heat-induced cells allows the polyme
rase to transcribe past this site. Therefore, the DRB-sensitive step i
s probably not rate-limiting for hsp70 transcription under uninduced c
onditions. DRB treatment did, however, lead to the reduction of KMnO4
hyper-reactivity on hsp70 and hsp26 in a region correlating with open
polymerase and/or early elongation complexes, suggesting a site for th
e DRB-sensitive polymerase modification step. Finally, we used the tec
hniques of polymerase-DNA cross-linking and KMnO4 hyper-reactive site
mapping to analyze the natural polymerase termination process at the 3
'-end of the hsp26 gene. The data obtained are consistent with polymer
ases terminating at multiple sites downstream of the polyadenylation s
ite.