FIG 6.

GAL10 antisense transcription is regulated by histone H2B ubiquitin conjugase and Paf1p. (A and B) Analysis of GAL10 antisense RNA in the Δrad6 and wild-type strains. (C) ChIP analysis of Myc-tagged Rad6p at the 5′ and 3′ ends of the GAL10 coding sequence, Chr.-V, and 5′ ends of the ADH1 and GAL7 coding sequences. (D) ChIP analysis of Flag-tagged histone H2B and HA-tagged ubiquitin at the 5′ and 3′ ends of the GAL10 coding sequence in the absence of the RING domain of Bre1p (the bre1Δ500 strain without 500 amino acids at Bre1p's C terminus that contain the RING domain). Immunoprecipitation was carried out using anti-Flag and anti-HA antibodies against Flag-tagged histone H2B and HA-tagged ubiquitin as described previously (31). (E and F) ChDIP analysis for the levels of ubiquitylated histone H2B in the wild-type (YKH045) and bre1Δ500 strains expressing Flag-tagged H2B and HA-tagged ubiquitin. (G) Analysis of histone H4 acetylation and histone H3 levels at GAL10 in the wild-type and Δrad6 strains. (H) Analysis of Rpb1p association with the 3′ and 5′ ends of the GAL10 coding sequence in the wild-type and bre1Δ500 strains. The maximum ChIP signal was set to 100, and the other ChIP signals relative to maximum ChIP signal were plotted. (I and J) Analysis of GAL10 antisense RNA in the Δpaf1 and wild-type strains.