Figure 5. Polyribosome or RNP enrichment of Lhb or Dusp1 mRNA by GnRH is not affected by rapamycin.

A) Summary of three independent determinations of polyribosome or RNP enrichment of Lhb and Dusp1 mRNA by GnRH either alone or with 30 minute pretreatment with the mTOR inhibitor rapamycin showing that neither Lhb nor Dusp1 enrichment is sensitive to inhibition of cap-dependent translation. Phospho-4EBP is reduced by pretreatment with rapamycin and the hypo- and un-phosphorylated α, β and γ isoforms are increased in abundance. Chemiluminescent images are presented in inverted grayscale. C) Summary of three independent Western blot determinations of MAPK 1/3 phosphorylation by GnRH after pretreatment with vehicle or rapamycin showing no effect on basal or GnRH-induced levels of phospho-MAPK 1/3. D) Summary of three independent determination of EIF4E phosphorylation after pretreatment with vehicle or rapamycin showing a modest increase in EIF4E phosphorylation in response to GnRH in both vehicle and rapamycin-treated cells. In both C and D dashed lines represent control ratio of phospho-MAPK1/3 to total MAPK 1/3 or phospho-EIF4E to total-EIF4E normalized to one. Asterisks indicate significant difference from untreated control as determined by Student’s t test and groups with different letters are significantly different from each other as determined by ANOVA and post hoc analysis with Tukey’s HSD test.