<i>dNf1</i> systemic growth related RAS/ERK and cAMP/PKA signals appear functionally and topographically distinct.

<p>(A) The elevated larval CNS pERK level of <i>dNf1</i> mutants is reduced by neuronal expression of <i>dNf1</i>, but not by neuronal or heat-shock induced ubiquitous expression of PKA*. Western blot of pERK levels in larval CNS of the indicated genotypes. In lane 6, larvae received a daily 20 min 37°C heat shock throughout development, a protocol that suppresses the <i>dNf1</i> growth defect <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003958#pgen.1003958-The1" target="_blank">[4]</a>. (B) Structure of <i>UAS-PKA*</i> transgenes with 1 to 5 UAS elements. The lethality of these transgenes when driven with either <i>Ac5C-Gal4</i> or <i>elav-Gal4</i> is indicated by † whereas (−) indicates viable offspring. (C) Western blot of adult head lysates showing relative expression of <i>GMR-Gal4</i>-driven transgenic PKA*. Tubulin is used as a loading control. (D) Expression of PKA* or knockdown of <i>dnc</i> by shRNAi in the ring gland rescues the <i>dNf1</i> pupal size defect. In contrast, <i>UAS-dNf1</i> expression with the same ring gland drivers fails to restore systemic growth. (E–H) Expression pattern of <i>Akh-Gal4</i> driving <i>UAS-GFP</i>, co-stained with DAPI and anti-dNF1. GFP expression in the corpora cardiaca (CC) is indicated. Scale bar = 50 µm. As previously noted <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003958#pgen.1003958-Walker2" target="_blank">[74]</a>, anti-dNf1 staining is strong in the CNS, whereas staining in the ring gland is close to background.