17β-estradiol-induced regulation of the novel 5-HT1A-related transcription factors NUDR and Freud-1 in SH SY5Y cells

Abstract

Nuclear deformed epidermal autoregulatory factor-1 (NUDR/Deaf-1) and five prime repressor element under dual repression (Freud-1) are novel transcriptional regulators of the 5-HT(1A) receptor, a receptor that has been implicated in the pathophysiology of various psychiatric illnesses. The antidepressant effect of 17β-Estradiol (17βE(2)) is purported to involve the downregulation of this receptor. We investigated the possible role of NUDR and Freud-1 in 17βE(2)-induced downregulation of the 5-HT(1A) receptor in the neuroblastoma cell line SH SY5Y. Cells were treated with 10 nM of 17βE(2) for 3 or 48 h, followed by a 24-h withdrawal period. Proteins were isolated and analyzed by western blotting. 17βE(2) treatment increased NUDR immunoreactivity while Freud-1 and the 5-HT(1A) receptor showed significant decreases. Upon withdrawal of 17βE(2), protein expression returned to control levels, except for NUDR, which remained significantly elevated in the 3-h treatment. Taken together, these data support a non-genomic downregulation of 5-HT(1A) receptor protein by 17βE(2), which does not involve NUDR and Freud-1. Rather, changes in both transcription factors seem to be compensatory/homeostatic responses to changes in 5-HT(1A) receptor induced by 17βE(2). These observations further highlight the importance of NUDR and Freud-1 in regulating 5-HT(1A) receptor expression.

Fig. 1

NUDR protein levels in SH-SY5Y cells after 3 h (a) and 48 h (b). Estradiol treatment (TX) and after 24-h withdrawal (TW). Data represent mean ROD ± SEM where sample size (N) for each treatment group (control, TX, and TW) is 3. ANOVA p values for 3 and 48 h are 0.0020, 0.0111, respectively. Post-hoc Dunnet’s test (TX or TW vs. control), * p < 0.05, ** p < 0.01

Fig. 2

Freud-1 protein levels in SH-SY5Y cells after 3 h (a) and 48 h (b). Estradiol treatment (TX) and after 24 h withdrawal (TW). Data represent mean ROD ± SEM where sample size (N) for each treatment group (control, TX, and TW) is 3. ANOVA p values for 3 and 48 h are 0.0294, 0.0288, respectively. Post-hoc Dunnet’s test (TX and TW vs. control), * p < 0.05, ** p < 0.01

Fig. 3

5-HT1_A receptor protein levels in SH-SY5Y cells after 3 h (a) and 48 h (b). Estradiol treatment (TX) and after 24-h withdrawal (TW). Data represent mean ROD ± SEM where sample size (N) for each treatment group (control, TX, and TW) is 3. ANOVA p values for 3 and 48 h are 0.0007 and 0.0222, respectively. Post-hoc Dunnett’s test (TX and TW vs. control), * p < 0.05, ** p < 0.01