TY - JOUR TI - Characterisation of the biflavonoid hinokiflavone as a pre-mRNA splicing modulator that inhibits SENP AU - Pawellek, Andrea AU - Ryder, Ursula AU - Tammsalu, Triin AU - King, Lewis J AU - Kreinin, Helmi AU - Ly, Tony AU - Hay, Ronald T AU - Hartley, Richard C AU - Lamond, Angus I A2 - Valcárcel, Juan VL - 6 PY - 2017 DA - 2017/09/08 SP - e27402 C1 - eLife 2017;6:e27402 DO - 10.7554/eLife.27402 UR - https://doi.org/10.7554/eLife.27402 AB - We have identified the plant biflavonoid hinokiflavone as an inhibitor of splicing in vitro and modulator of alternative splicing in cells. Chemical synthesis confirms hinokiflavone is the active molecule. Hinokiflavone inhibits splicing in vitro by blocking spliceosome assembly, preventing formation of the B complex. Cells treated with hinokiflavone show altered subnuclear organization specifically of splicing factors required for A complex formation, which relocalize together with SUMO1 and SUMO2 into enlarged nuclear speckles containing polyadenylated RNA. Hinokiflavone increases protein SUMOylation levels, both in in vitro splicing reactions and in cells. Hinokiflavone also inhibited a purified, E. coli expressed SUMO protease, SENP1, in vitro, indicating the increase in SUMOylated proteins results primarily from inhibition of de-SUMOylation. Using a quantitative proteomics assay we identified many SUMO2 sites whose levels increased in cells following hinokiflavone treatment, with the major targets including six proteins that are components of the U2 snRNP and required for A complex formation. KW - hinokiflavone KW - RNA splicing KW - SUMO KW - SENP1 KW - U2 snRNP JF - eLife SN - 2050-084X PB - eLife Sciences Publications, Ltd ER -