Fig. 2

Removal of the N-terminal region of SETD2 leads to its stabilization. a, d Western blot of whole-cell lysates probed with the depicted antibodies. Lysates of 293T cells expressing Halo-ySet2 constructs were prepared after 12 h of MG132 (10 µM) treatment. U-untransfected. b Cartoon illustrating the known domains of yeast Set2 (ySet2), SETD2, and the SETD2 C-terminus region that shares the conserved domains with ySet2. c Microscopy images showing the expression and localization of GFP-SETD2 C and ySet2. The scale bar is 1 mm for 10 × images and 10 µm for the magnified ones. e RNA was isolated from transfected cells described in d and RT-PCR was performed to check the SETD2 transcript levels. GAPDH was used as a normalization control. SETD2_1 primer pair binds to the N-terminal segment of SETD2 transcripts, whereas, SETD2_2 binds to the C-terminal region which is common in both SETD2 FL and C. f Cartoon illustrating the SETD2 segments used in the cycloheximide-chase experiment. g Microscopy images showing the expression and localization of the GFP-SETD2 N + Catalytic Domain of SETD2. The scale bar is 10 µm. h Western blot of whole-cell lysates probed with the depicted antibodies. Lysates of 293T cells expressing Halo-FLAG-SETD2 constructs were prepared after cycloheximide (10 µg/ml) treatment. The duration of the treatment is shown in the figure. HE- High Exposure, LE-Low Exposure