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. 2016 May;57(5):868–881. doi: 10.1194/jlr.M067447

Fig. 2.

Fig. 2.

ATRA induction of nSMase2 is at the transcriptional level. A: MCF7 cells were seeded in 60 mm dishes and stimulated with vehicle (DMSO) or ATRA (1 μM) for 12 h. At this time, transcription was interrupted with actinomycin D (ActD; 1 μg/ml) for 0, 3, 6, and 9 h as shown. At these times, RNA was extracted and nSMase2 expression analyzed by qRT-PCR (P > 0.1; n = 6). B: MCF7 cells were seeded in 60 mm dishes and, 24 h later, were cotransfected with 1 μg nSMase2 promoter-luciferase and 0.5 μg pEF6-LacZ for 6 h prior to stimulation with vehicle (DMSO) or ATRA (1 μM) for 24 h. Cells were lysed, and luciferase and galactosidase activity were determined as described in Materials and Methods (* P < 0.05 vs. vehicle, n ≥ 3). C: MCF7 cells were seeded in 100 mm dishes and stimulated with vehicle (DMSO) or ATRA (1 μM) for 24 h. hnRNA levels were analyzed as described in Materials and Methods (** P < 0.01, *** P < 0.001 vs. vehicle, n = 3).