Tasor

The following antibodies were used: AFB1 (Santa Cruz, no. sc-393403), SAFB2 (Santa Cruz, no. sc-514963), SAFB1/2 (HET) (human: Merck/Sigma-Aldrich, no. sc05-588; mouse: LSBio, no. LS-C2886411), SLTM (Invitrogen, no. PA5-59154), ORF1p (human: abcam, no. ab230966; mouse: abcam, no. ab216324), TASOR (Sigma-Aldrich, no. HPA006735), 1H4 (p-SR) (Merck/Sigma-Aldrich, no. MABE50), RBM12B (Bethyl, no. A305-871A-T), RBMX (Cell Signaling Technology, no. 14794 S), NCOA5 (Bethyl, no. A300-790A-T), ZNF638 (Sigma-Aldrich, no. ZRB1186), ZNF326 (Santa Cruz, no. sc-390606), TRA2B (Bethyl, no. A305-011A-M), U2AF2 (U2AF65; Santa Cruz, no. sc-53942), TUBULIN (Santa Cruz, no. sc-32293), SRRM1 (abcam, no. ab221061), SRRM2 (SC35) (Sigma-Aldrich, no. S4045), SON (Sigma-Aldrich, no. HPA023535), DHX9 (abcam, no. ab183731), U1-70K (SySy, no. 203011), PRP8 (Santa Cruz, no. sc-55533), RNAPII (Creative Biolabs, no. CBMAB-XB0938-YC), IgG normal mouse (Santa Cruz, no. sc-2025), SRSF1 (Santa Cruz, no. sc-33652), SRSF2 (abcam, no. ab204916), SRSF3 (Elabscience, no. E-AB-32966), SRSF7 (MBL, no. RN079PW), RB1 (Cell Signaling Technology, no. 9309 S), TRA2B (Santa Cruz, no. sc-166829) and YTHDC1 (Proteintech, no. 14392-1-AP).

Cells were lysed in radioimmunoprecipitation assay (RIPA) lysis and extraction buffer containing protease inhibitor for 20 min. The lysate was clarified by centrifugation at 4°C for 10 min at 12,000 rpm. The samples were boiled at 95°C in SDS sample buffer containing β-mercaptoethanol and resolved by SDS-PAGE electrophoresis. The resolved proteins were transferred to a polyvinylidene difluoride (PVDF) membrane using the Trans-Blot Turbo semidry transfer system (Bio-Rad), and the membrane was blocked in 1× PBS containing 0.1% Tween 20 and 5% skim milk powder for 1 h at room temperature. The membranes were probed with specific primary and secondary antibodies. The primary antibodies used were mouse anti-actin (catalog no. ab3280) and PPHLN1 (catalog no. ab69569), which were purchased from Abcam. SETDB1 (catalog no. 11231-1-AP) and MPP8 (catalog no. 16796-1-AP) were purchased from Proteintech. ZNF638/NP220 (catalog no. A301-548A) was purchased from Bethyl Laboratories; TASOR (catalog no. HPA006735) was from Sigma-Aldrich. Goat anti-rabbit-horseradish peroxidase (HRP) (catalog no. 111-035-003) was obtained from Jackson ImmunoResearch. Goat anti-mouse-HRP (catalog no. 32430) was obtained from Thermo Fisher.