A hyperphosphorylated form of RNA polymerase II is the major interphase antigen of the phosphoprotein antibody MPM-2 and interacts with the peptidyl-prolyl isomerase Pin1

The monoclonal antibody MPM-2 recognizes a subset of M phase phosphoprotein s in a phosphorylation-dependent manner. It is believed that phosphorylatio n at MPM-2 antigenic sites could regulate mitotic events since most of the MPM-2 antigens identified to dale have N1 phase functions. In addition, man y of these proteins are substrates of the mitotic regulator Pin1, a peptidy l-prolyl isomerase which is present throughout the cell cycle and which is thought to alter its mitotic targets by changing their conformation. In int erphase cells, most MPM-2 reactivity is confined to nuclear speckles, We re port here that a hyperphosphorylated form of the RNA polymerase II largest subunit is the major MPM-2 interphase antigen, These findings were made pos sible by the availability of another monoclonal antibody, CC-3, that was pr eviously used to identify a 255 kDa nuclear matrix protein associated with spliceosomal components as a hyperphosphorylated form of the RNA polymerase II largest subunit, MPM-2 recognizes a phosphoepitope of the large subunit that becomes hyperphosphorylated upon heat shock in contrast to the phosph oepitope defined by CC-3, whose reactivity is diminished by the heat treatm ent. Therefore, these two antibodies may discriminate between distinct func tional forms of RNA polymerase II. We also show that RNA polymerase II larg e subunit interacts with Pin1 in HeLa cells, Pin1 may thus regulate transcr iptional and post-transcriptional events by catalyzing phosphorylation-depe ndent conformational changes of the large RNA polymerase II subunit.