<i>Ce-age-1</i> is expressed in amphidial neurons and other tissues.

<p>Fluorescence (A,C) and DIC (B,D) images of transgenic <i>C. elegans</i> first-stage larvae expressing <i>Ce-age-1p::Ce-age-1(102bp) ::egfp::Ce-age-1t</i> from an extra-chromosomal array. (A,B) Strong expression of the EGFP reporter was present in amphidial neurons (a), a neuron or support cell anterior to the nerve ring (c), and the sphincter connecting the pharynx to the intestine (s). Weak expression was present in the intestine (i), hypodermis (h), and a phasmidial neuron (p). (C,D) EGFP reporter expression was present in the amphidial neurons AWC (short arrow) and ASJ (long arrow). Cell bodies of the amphidial neurons align just lateral to the black lines in panel D <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038587#pone.0038587-Srinivasan1" target="_blank">[74]</a>. Scale bars = 100 µm.</p

<i>Ss-age-1</i> is expressed in amphidial neurons, the intestine, and other tissues.

<p>Fluorescence (A,C) and DIC (B,D) images of transgenic <i>S. stercoralis</i> post-free-living first-stage larvae expressing <i>Ss-age-1p::egfp::Ss-era-1t</i> from an extra-chromosomal array. (A,B) Expression of the EGFP reporter was present in the intestine (i), gonadal primordium (g), amphidial/head neuron (a), hypodermis (h), and phasmidial/tail neuron (p). (C,D) Expression of the EGFP reporter was present in an amphidial neuron (long arrow), with positional homology to AWC in <i>C. elegans</i>. The other cell body of the amphidial neuron pair is out of the plane of focus (short arrow). Cell bodies of the amphidial neurons align just lateral to the black lines in panel D <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038587#pone.0038587-Srinivasan1" target="_blank">[74]</a>. Scale bars = 100 µm.</p

Sites of <i>Ss-age-1</i> expression in transgenic <i>S. stercoralis</i> post-free-living first-stage larvae.

<p>Sites of <i>Ss-age-1</i> expression in transgenic <i>S. stercoralis</i> post-free-living first-stage larvae.</p

<i>Ss</i>-AGE-1 is a homolog of the <i>Ce</i>-AGE-1 PI3 kinase catalytic subunit.

<p>(A) Intron-exon structure of the <i>Ss-age-1</i> unspliced mRNA sequence. Grey boxes indicate the three exons, with the numbers above indicating the first and last base pairs of the exon. Introns are indicated by slanted lines between the exons, with the numbers indicating the total intron length. The 5′ and 3′ untranslated regions (UTR) are indicated with horizontal lines, with the italicized numbers indicating the total length of the UTR in base-pairs (bp). (B) Domain structure of the <i>Ss</i>-AGE-1 predicted protein. Shaded boxes represent the five protein family domains, with the numbers indicating the first and last amino acids of the domain. (C) Phylogenetic analysis of <i>Ss</i>-AGE-1. The predicted <i>Ss</i>-AGE-1 protein groups with other class I PI3 kinase catalytic subunits, including <i>Ce</i>-AGE-1. Abbreviations: <i>Strongyloides stercoralis</i> (Ss), <i>Strongyloides ratti</i> (Sr), <i>Parastrongyloides trichosuri</i> (Pt), <i>Brugia malayi</i> (Bm), <i>Caenorhabditis briggsae</i> (Cb), <i>Caenorhabditis elegans</i> (Ce), <i>Homo sapiens</i> (Hs), <i>Drosophila melanogaster</i> (Dm), and <i>Saccharomyces cerevisiae</i> (Sc). Accession numbers listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038587#s2" target="_blank">Methods</a>.</p

<i>S. stercoralis</i> L3i activation is attenuated by the PI3 kinase inhibitor LY294002.

<p><i>In vitro</i> activation of <i>S. stercoralis</i> L3i under host-like culture conditions by incubation in DMEM, 10% canine serum, and 12.5 mM reduced glutathione for 24 hours at 37°C and 5% CO₂. The percentage of L3i that resumed feeding, a hallmark of activation, was scored by ingestion of FITC into the pharynx. Conditions included DMSO (carrier) positive control and M9 buffer negative control. The PI3 kinase inhibitor LY294002 was evaluated at 100 µM, 50 µM, and 10 µM, with each condition compared to the DMSO control using a logistic regression analysis. Error bars represent +1 SEM and parenthetical integers show the total number of L3i evaluated for each condition.</p