TRANSCRIPTION-DEPENDENT REDISTRIBUTION OF THE LARGE SUBUNIT OF RNA-POLYMERASE-II TO DISCRETE NUCLEAR DOMAINS

A subpopulation of the largest subunit of RNA polymerase II (pol II LS ) is located in 20-50 discrete subnuclear domains that are closely lin ked to speckle domains, which store splicing proteins. The speckle-ass ociated fraction of Pol II LS is hyperphosphorylated on the COOH-termi nal domain (CTD), and it is highly resistant to extraction by detergen ts. A diffuse nucleoplasmic fraction of pol II LS is relatively hypoph osphorylated on the CTD, and it is easily extracted by detergents. In transcriptionally active nuclei, speckle bound hyperphosphorylated Pol II LS molecules are distributed in irregularly shaped speckle domains , which appear to be interconnected via a reticular network. When tran scription is inhibited, hyperphosphorylated pol II LS and splicing pro tein SC35 accumulate in speckle domains, which are transformed into en larged, dot-like structures lacking interconnections. When cells are r eleased from transcriptional inhibition, Pol IIO and SC35 redistribute back to the interconnected speckle pattern of transcriptionally activ e cells. The redistribution of pol II and SC35 is synchronous, reversi ble, and temperature dependent. It is concluded that: (a) hyperphospho rylation of pol II LS's CTD is a better indicator of its tight associa tion to discrete subnuclear domains than its transcriptional activity; (b) during states of transcriptional inhibition, hyperphosphorylated pol II LS can be stored in enlarged speckle domains, which under the l ight microscope appear to coincide with the storage sites for splicing proteins; and (c) pol II and splicing proteins redistribute simultane ously according to the overall transcriptional activity of the nucleus .