Newly identified loci display phenotypes resembling DA deficiency.

<p>(A) Enhancement of <i>daf-36(k114)</i> gonadal Mig defects is seen upon knockdown of <i>daf-9</i>, <i>dhs-16</i>, and <i>emb-8</i>. Arrowheads indicate the arms of the gonad, visible along the body due to failure of proper distal tip cell migration. (B) Mig and Daf-c phenotypes of <i>dhs-16(tm1890)</i> deletion mutants, under conditions of cholesterol deprivation (upper image) and 27°C (lower image). (C) Analysis of dauer formation at 27°C on NGM. <i>dhs-16(tm1890)</i> mutants show Daf-c phenotypes similar to <i>daf-36(k114)</i> (<i>N</i> = 3, M±SD; **<i>p</i><0.01). (D) Analysis of the gonadal Mig defects of <i>dhs-16</i> null animals on NGM without added cholesterol (<i>N</i> = 3, M ± SD; **<i>p</i><0.01, *<i>p</i><0.05). (E) RNAi knockdown of <i>dhs-16</i> and <i>emb-8</i> in wild-type worms at 27°C induces Daf-c phenotypes, similar to knockdown of <i>daf-36</i> (<i>N</i>≥4, M ± SD; **<i>p</i><0.005). (F) Genetic epistasis analysis of <i>dhs-16(tm1890)</i> (Daf-c) together with Daf-d mutations in transcription factors of insulin/IGF, TGF-β, and DA signaling show that DHS-16 works downstream of DAF-16/FOXO and DAF-5/SKI, but upstream of DAF-12/NHR (<i>N</i> = 3, M ± SD; **<i>p</i><0.01). (G) Similar genetic epistasis analysis of <i>emb-8</i> RNAi-induced dauer phenotypes at 27°C suggests that EMB-8 acts downstream of DAF-16 and upstream of DAF-12 (<i>N</i> = 3, M ± SD; **<i>p</i><0.01). (H) The DAF-12 target gene <i>mir-241</i>, a <i>let-7</i> related microRNA, shows reduced expression in <i>dhs-16</i> mutants under low cholesterol conditions at 25°C (<i>N</i> = 3, M ± SD; **<i>p</i><0.01, *<i>p</i><0.05).