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Fig. 2. LAG-1 and Fos/Jun cis binding elements are required for egl-13 expression at cell specification and for rescue of egl-13 mutant defects. (A) Conservation of transcription factor binding sites within the 1.3 kb enhancer of egl-13. Conserved cis-elements are shown in bold. Dark gray boxes represent sequences sharing remarkable identity with C. briggsae: 106 bp box 1 in elegans shares 88.2% identity with briggsae, 21 bp box 2 shares 100%, 65 bp box 3 shares 89.6%, 36 bp box 4 shares 94.4%, 142 bp box 5 shares 91.7%, and 45 bp box 6 shares 93.5%. Trans factor abbreviations (in order of relative upstream positions): Hb, Hunchback; Dof, Dof domain; MBF1, Multi-protein binding factor; Y-box, Y-box binding protein; Etsr, ETS domain related; HOM, homeodomain; POU, POU domain; HES, Hairy/Enhancer of split; TFIID, transcription factor II D; ETS domain; NBF, Nonamer binding protein; STRE, stress response element; ZnF, zinc finger; LIM, LIM domain; NF-B or Rel homology; Sp1 or ZnF; FOX, F-box/Forkhead; GATA, GATA binding factor; BR-C, Broad complex or ZnF; E2F factor; TBP, TATA binding protein; SRY, SRY or high mobility group (HMG). (B) Effect of LAG-1 or Fos/Jun deletion on temporal uterine egl-13^FL::GFP fluorescence. DIC images showing medial uterine-vulval development. Lateral expression of egl-13^FL::GFP in cells is seen here just after the cells have divided. Neither egl-13^FLLAG-1::GFP nor egl-13^FLFos/Jun::GFP transgenic animals fluoresce at specification. egl-13^FLLAG-1::GFP expresses in all descendants later, whereas later expression of egl-13^FLFos/Jun::GFP is only in uv1. Brackets encompass the cells. Dashed ovals surround uterine area where fluorescence is lost. Asterisks mark body wall expression. Scale bar: 10 µm. (C) Four constructs of egl-13^FL::cDNA were generated with intact, singly, or doubly-deleted LAG-1 and Fos/Jun binding sites. L4 stage hermaphrodites from independent egl-13(ku194) transgenic lines for each construct were scored over 4 days for egg laying. Egl+ indicates healthy appearance and egg laying. Egl+/-indicates egg laying but bloated appearance and/or rupture from internal hatching. Egl-indicates no egg laying and eventual rupture. ku194 is 100% Egl-(n=100s, not shown). The strain designations are provided in Materials and methods.

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