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. 2020 Feb 21;6(8):eaay5154. doi: 10.1126/sciadv.aay5154

Fig. 2. Light controls the effects of opto-pomalidomide in mediating IKZF1/3 degradation.

Fig. 2

(A) A schematic diagram showing that UVA irradiation activates opto-pomalidomide in cell culture. (B) UVA irradiation activates opto-pomalidomide to mediate the interaction between CRBN and IKZF1. IB analysis of whole-cell lysis (WCL) and Flag–immunoprecipitation (IP) derived from HEK293T cells transfected with indicated plasmids in the presence of pomalidomide or opto-pomalidomide with/without UVA irradiation (365 nm) for 15 min. Cells were treated with 10 μM MG132 for 12 hours before harvest. (C) UVA irradiation activates opto-pomalidomide to mediate the ubiquitination of IKZF1 by CRBN in cells. IB analysis of WCL and Ni–nitrilotriacetic acid (NTA) pull down products derived from HEK293T cells transfected with indicated plasmids in the presence of pomalidomide or opto-pomalidomide with or without UVA irradiation (365 nm) for 15 min. Cells were treated with 10 μM MG132 for 12 hours before harvest. (D) Opto-pomalidomide does not promote the degradation of IKZF1/3 without UVA irradiation. IB analysis of WCL derived from MM.1SCRBN+/+ versus MM.1SCRBN−/− cells in the presence of pomalidomide or opto-pomalidomide for 12 hours. (E) UVA irradiation activates opto-pomalidomide to promote the degradation of IKZF1/3 in cells. IB analysis of WCL derived from MM.1S cells in the presence of opto-pomalidomide with UVA irradiation (365 nm) as indicated time. (F) UVA irradiation–activated opto-pomalidomide inhibits MM.1S cell proliferation in a dose-dependent manner. MM.1S cells were treated by pomalidomide versus opto-pomalidomide with or without UVA irradiation (365 nm) for 15 min and then subjected to CCK-8 cell viability assay. (G) Pomalidomide reduces MM.1S cell proliferation in a CRBN-dependent manner. MM.1SCRBN+/+ and MM.1SCRBN−/− cells were treated by pomalidomide for 72 hours and then subjected to CCK-8 cell viability assay.