Figure S1. DAF-16 regulates
sod-3 and
lys-7 cell-autonomously, and an
mtl-1 translational but not transcriptional reporter responds well to
daf-2 inhibition (Related to Figure 1). (A) We reproduced our previous report of cell-autonomous regulation of
sod-3 by DAF- 16 (Libina et al., 2003), using (integrated) transgenic strains that are
daf-16(+) in only one tissue (intestine, muscles, neurons). Top panels: Psod-3::gfp (cytoplasmic) is expressed mainly in the pharynx (ph), muscles (m) and neurons (nerve ring) (n) of wild type, and is up-regulated in most tissues of
daf-2(-) mutants (inset, full exposure time) in a
daf-16-dependent manner. 'h', hypodermis. Bottom panels: intestinal DAF-16 upregulates Psod-3::gfp only in the intestine (inset, full exposure time). Muscle and neuronal DAF-16 up-regulate Psod-3::gfp only in muscles and neurons, respectively. Note that the muscle-
daf-16(+) construct contains a
rol-6 co-injection marker, which twists the animal. Young adults, 250X magnification. Scale bar: 50 um. Representative images from 2 or more experiments are shown for all figures. 2 (B) DAF-16 regulates
lys-7 cell-autonomously. Top panels: nuclear-localized Plys-7::rfp is expressed mainly in the intestine (i) of wild type, and is up-regulated in
daf-2(-) mutants in a
daf-16-dependent manner. The body is outlined ('ph', pharynx). Bottom panels: intestinal, but not muscle or neuronal (not shown) DAF-16 up-regulates Plys- 7::rfp in the intestine. Bottom panel (left, inset): gfp::
daf-16 expression (green). Young adults, 250X magnification. Scale bar: 50 um. Bright field (bottom middle) and fluorescence (RGB channels on the Axioplan 2 compound microscope) (bottom right) pictures of the same animal at higher magnification are shown. Arrowheads indicate two green (GFP::DAF-16) and red (Plys-7::rfp) intestinal cells. Bottom panel (right, inset): Plys-7::rfp expression (red). Note that cells expressing higher levels of gfp::
daf-16 express higher levels of Plys-7::rfp. 400X magnification. Scale bar: 32 um. (C) A translational, but not a transcriptional reporter of
mtl-1 responded well to
daf-2 inhibition. Left two panels: nuclear localized Pmtl-1::rfp is expressed mainly in the intestine (i) of wild type, and showed modest response to
daf-2 knockdown. Right panels: an
mtl-1 translational fusion, which is not nuclear localized, is induced robustly in the intestine upon
daf-2 inhibition. The body is outlined ('ph', pharynx). Young adults, 100X magnifications. Scale bar: 130 um.