FIG 6.

Opposite effects of Homer1a- and Homer1b/c-specific knockdown on GnRH-induced gonadotropin β-subunit gene expression. On day 1, cells were transfected with either scrambled (Ctrl), Homer1a (H1a), or Homer1b/c (H1bc) siRNA for 48 h. On day 2, cells were serum starved overnight. On day 3, cells were stimulated with 5 nM GnRH for the indicated times. (A, B, and E) Knockdown efficiency and siRNA specificity of Homer1a and Homer1b/c siRNAs on mRNA level (A and B) and on protein level (E). (C and D) Knockdown of Homer1b/c upregulates GnRH-mediated FSHβ and LHβ gene expression, while knockdown of Homer1a reduces FSHβ gene expression but does not alter LHβ. For RT-qPCR, the data shown are means ± SEM of triplicate samples from one experiment and are representative of three independent experiments. For Western blots, a representative figure is shown and densitometry is based on three independent experiments. Two-way ANOVA with Bonferroni posttest corrections; *, P < 0.05; **, P < 0.01.