THE carboxy-terminal domain (CTD) of the large subunit of RNA polymera
se II is essential in vivo(1-4), and is found in either an unphosphory
lated (IIa) or hyperphosphorylated (IIo) form(5,6). The Drosophila uni
nduced hsp70 and hsp26 genes, and the constitutively expressed beta-1
tubulin and Gapdh-2 genes, contain an RNA polymerase II complex which
pauses after synthesizing a short transcript(7-10). We report here tha
t, using an in vivo ultraviolet crosslinking technique(11) and antibod
ies directed against the IIa and IIo forms of the CTD12, these paused
polymerases have an unphosphorylated CTD. For genes containing a 5' pa
used polymerase, passage of the paused RNA polymerase into an elongati
onally competent mode in vivo coincides with phosphorylation of the CT
D. Also, the level of phosphorylation of the CTD of elongating polymer
ases is shown not to be related to the level of transcription, but is
promoter specific.