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. Author manuscript; available in PMC: 2020 Jun 17.
Published in final edited form as: Curr Biol. 2019 May 30;29(12):1954–1962.e4. doi: 10.1016/j.cub.2019.04.073

Figure 5. The N-terminus of Cry2 interacts with Det1 to stabilize Cop1’s substrates and inhibit GR activity.

Figure 5.

(A) HA-tagged Det1 was co-expressed in HEK-293T cells with either FLAG-tagged wild-type Cry2 or FLAG-tagged Cry2 truncation mutants followed by immunoprecipitation from cell extracts with an anti-HA resin. Cells extracts (CE) and immunoprecipitates (IP) were analyzed by immunoblotting. Roman numbers indicate different exposure times: I, short exposure; II, long exposure.

(B) Schematic representation of wild-type Cry2 and Cry2 truncation mutants.

(C) FLAG-tagged Cry2 or FLAG-tagged Cry2-N130 truncation mutant were expressed in HEK-293T cells and immunoprecipitated from cell extracts with an anti-FLAG resin. CEs and immunoprecipitates were then analyzed by immunoblotting.

(D) U-2OS cells were transfected with either an empty vector (EV) or with a vector expressing Cry2-N130. 18 hours after transfection, cells were treated with CHX for the indicated times and cell extracts were immunoblotted.

(E) qPCR analysis of cDNAs prepared from U-2OS cells infected with lentiviruses expressing Cry2-N130 under the control of a doxycycline-inducible promoter and treated with dexamethasone where indicated. See also Figures S3, S4 and S5.