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This study examines the impact of increasing concentrations of NBBS on glucocorticoid, thyroid hormone, and progesterone receptor transactivation in CV1 cells lacking endogenous steroid receptors. Specifically, it was found that NBBS does not affect transactivation mediated by human GRα and TRβ, while it inhibits the action of the human progesterone receptor A-form. Nuclear receptor transactivation assays were conducted alongside Western blot analysis to evaluate the stability of the androgen receptor post-NBBS treatment. Results highlight the selective influence of NBBS on steroid hormone signaling.
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no hormone 8 Glucocorticoid 10-11 M 6 RLU x104 4 2 - 100 10 1 NBBS [µM] human GRa Fig. 1S NBBS does not effect the transactivation mediated by the human GRα. CV1 cells lacking endogenous steroid receptors or TR were tested for the nuclear receptor transactivation assays as described in figure 4. Increasing concentrations of NBBS were tested for inhibtion of glucocorticoid- (dexamethason) mediated transactivation by transiently transfected cells with the hGRalpha expression vector. Fig. 1S
no hormone Thyroid hormone 10-6 M A 10 8 6 RLU x104 4 2 - NBBS [µM] 100 10 1 human TRβ Fig. 2S
no hormone Thyroid hormone 10-7 M B 25 20 15 RLU x104 10 5 - NBBS [µM] 100 10 1 human TRβ Fig. 2S
no hormone Thyroid hormone 10-8 M C 25 20 15 RLU x104 10 5 - NBBS [µM] 100 10 1 human TRβ Fig. 2S NBBS does not effect the transactivation mediated by the human TRβ. CV1 cells lacking endogenous steroid receptors or TR were tested for the nuclear receptor transactivation assays as described in figure 4. Increasing concentrations of NBBS were tested for inhibtion of indicated or thyroid hormone- (A-C) mediated transactivation by transiently transfected cells with the human TRbeta expression vector. Fig. 2S
no hormone Progesterone 10-9 M 5 4 3 RLU x103 2 1 - NBBS [µM] 100 human PR A-form Fig. 3S NBBS inhibits the transactivation mediated by the human progesterone receptor A-form CV1 cells lacking endogenous steroid receptors were tested for the nuclear receptor transactivation assays as described in figure 4. The human PR-A lacks sequences from the N-terminus of the B-form due to an internal translation start site. Fig. 3S
10-5M NBBS 10-5M NBBS 10-4M NBBS 10-4M NBBS Control Control 3 days 7 days WB: a-AR Coomassie staining Fig. 4S NBBS does not influence AR stability. LNCaP cells were treated with 100 µM NBBS for three or seven days. Western blot was performed with anti-AR antibody. Coomassie staining was employed for equal protein loading. Fig. 4S
