Extended Data Fig. 8: SF2 binds to DDX5 and regulates IL-36R splicing and inflammatory responses in keratinocytes.
a, Mass spectrum analysis of DDX5-binding splicing factors in NHEKs. b, Co-immunoprecipitation analysis of the interaction of DDX5 and SF2. c, Immunoblot of HA-tagged or Flag-tagged proteins that bind to IL36R pre-mRNA by Flag-tagged antibody or HA-tagged antibody. d, RT-qPCR of CCL20, CXCL1, CXCL6, CCL17, CCL3, CCL11 and CCL27 in NHEKs in response to 100 ng/mL IL-36γ before and after SF2 was silenced. e, SF2-binding ESEs on exon 2, exon 3 and exon 4 of human IL1RL2 gene analyzed by the ESEfinder program (http://exon.cshl.edu/ESE). f, Schematic diagram represents IL36R pre-mRNA splicing regulated by SF2. Strong interaction of SF2 on the flanking exon 4 is responsible for skipping of the alternative exon 3 to generate sIL-36R. When the interaction of SF2 on exon 4 is weakened, either by ESE mutation in exon 4 or SF2 depletion, the alternative exon 3 is selected for IL-36R generation. g, Immunoblot of IL36R pre-mRNA-binding SF2 in nuclear extracts prepared from SF2–/– HeLa cells by Flag-tag antibody, in which IL36R reporter minigene or IL36R reporter minigene containing ESE mutation in Exon 4 was co-transfected with exogenous Flag-tagged SF2. Data represent at least three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001. P values were determined by two-way ANOVA. Data are presented as mean ± s.e.m.
