Figure 1. Endogenous GFP reporters provide precise and quantitative information on
gpdh-1 expression and localization:A) Design strategy for the endogenous
gpdh-1 reporters. These consist on the substitution of the whole
gpdh-1 coding sequence (CDS) to generate
gpdh-1(
dr153) (GFP::NLS),
gpdh-1(
dr154) (GFP), or
gpdh-1(
dr157) (GFP-
his-58). B) Images of
day1 adult animals expressing the indicated
gpdh-1reporter under normal (50 mM NaCl) and hypertonic (250 mM NaCl) conditions. The drIs4 reporter is shown as reference. All images were taken using the same exposure settings. Insets in
dr157 and
dr153 are longer exposures and show that these reporters express at low levels and give rise to nuclear enriched GFP signal in the hypodermis and intestine under hypertonic conditions. C) Relative fold induction of GFP intensity by hypertonic conditions (250 mM NaCl) for the reporters shown in B. Data is represented as mean ± S.D. ****p<0.0001, ns: nonsignificant. (Mann-Whitney test). Data were normalized to the mean expression levels at 50 mM NaCl for each genotype. D) Kinetics of
gpdh-1(
dr154) and drIs4 activation measured as the change in normalized GFP/TOF within 0, 2, 8 and 24h. Data was fit to a four parameter logistic (4PL) curve. N= 800-3000 for each data point. E) Images of
osm-7(
n1515);
gpdh-1(
dr154) or
osm-8(
n1518);
gpdh-1(
dr154) day 1 adults following 2 generation exposure to
ptr-23(RNAi). All images were taken using the same exposure time. F) COPAS quantification of the relative fold changes of GFP intensity of
osm-7(
n1515);
gpdh-1(
dr154) and
osm-8(
n1518);
gpdh-1(
dr154) exposed to either empty vector (EV) or
ptr-23(RNAi). Data for each genotype is normalized to the EV mean and represented as mean ± S.D. **p<0.005, ns: nonsignificant. (Mann-Whitney test). N=81-139 for each condition.