SETD2 is necessary to maintain H3K36me3 on HPV31 chromatin.

(A) Schematic of the HPV31 genome showing the location of the primer pairs utilized for ChIP analysis. (B, C) Chromatin and protein were harvested from CIN612 cells transduced with either control shRNA or SETD2 shRNA #2 for 72hr (T0), as well as after differentiation in high calcium medium for 72hr. ChIP was performed using an antibody to (B) H3K36me3 or (C) H3.1 using the indicated primer pairs across the HPV31 genome (primer sequences listed in S1 Table). Data of ChIP signals from three independent experiments were normalized to 1% of input used. Shown in the fold change in H3K36me3 or H3.1 binding relative to the first primer set, which is set to one. Error bars represent means ± standard error. Statistics were assayed using a student’s t test. * p≤ .05 and ** p≤ .01. (D) Western blot analysis was performed using antibodies to SETD2, to involucrin and K10 as differentiation controls and p84 as a loading control. Ca = calcium.