Anti-LEO1 (aa 140-240) polyclonal antibody

Parafibromin, the product of the HRPT2 (hyperparathyroidism-jaw tumor syndrome 2) tumor suppressor gene, is the human homologue of yeast Cdc73, part of the yeast RNA polymerase II/Paf1 complex known to be important for histone modification and connections to posttranscriptional events. By purifying cellular parafibromin and characterizing its associated proteins, we have identified a human counterpart to the yeast Paf1 complex including homologs of Leo1, Paf1, and Ctr9. Like the yeast complex, the parafibromin complex associates with the nonphosphorylated and Ser2 and Ser5 phosphorylated forms of the RNA polymerase II large subunit. Immunofluorescence experiments show that parafibromin is a nuclear protein. In addition, cotransfection data suggest that parafibromin can interact with a histone methyltransferase complex that methylates histone H3 on lysine 4. Some mutant forms of parafibromin lack association with hPaf1 complex members and with the histone methyltransferase complex, suggesting that disruption of these complexes may correlate with the oncogenic process.

The Paf1 complex (Paf1C) affects RNA polymerase II transcription by coordinating co-transcriptional chromatin modifications and helping recruit mRNA 3' end processing factors. Paf1C cross-links to transcribed genes, but not downstream of the cleavage and polyadenylation site, suggesting that it may interact with the nascent mRNA. Paf1C purified from Saccharomyces cerevisiae binds RNA in vitro, as do the purified Leo1 and Rtf1 subunits of the complex. In vivo cross-linking and immunoprecipitation of RNA associated with Paf1C (RNA-IP) show that Leo1, but not Rtf1, is necessary for the complex to bind RNA. Cells lacking Leo1 have reduced Paf1C recruitment as well as decreased levels of histone H3 and trimethylated H3 Lys(4) within transcribed chromatin. Together, these results suggest that association of Paf1C with RNA stabilizes its localization at actively transcribed regions where it influences chromatin structure.