Figures:
Figure 1: Survival and organismal phenotypes during
hypersalinity acclimation. a) Blood osmolality values for five
14-day acclimations after 24 hours at the target salinity (n=6 except at
75g/kg with n=4) and following extended exposure for 12 weeks (75g/kg,
n=8) and at MP (85/105g/kg, n=6). b) Body condition for the
same treatments as c . Letters in C and D were assigned using a
post-hoc Tukey test following one-way ANOVA.
Figure 2: Growth and feeding during 12-week exposure to
75g/kg salinity. a) Size comparison of smallest and largest
fish in the freshwater and 75g/kg treatment at the end of the
experiment. b) Weight gain for FW control and at 75g/kg, with
comparison between treatments. Significance was determined using
Kruskal-Wallis test by ranks with “ns”- not significant, “*”-
p<0.05, “**”- p<0.005, “***”-
p<0.0005. c) Percent of fish body weight consumed
daily over the course of the experiments, with weekly weighing
identified by dashed vertical lines.
Figure 3 : Salinity effects on the gill proteome for five
treatments. a) Volcano plots for each treatment with log2(Fold
Change) plotted against log10(p-value). n=8 for 85 and 105g/kg 14-day
acclimation and 75g/kg extended treatment, and n=6 for extended 85 and
105g/kg treatments. Significantly upregulated proteins are plotted in
red with the total number of sig. upregulated proteins labelled, and
significantly downregulated proteins are in blue. b) Overlap in
all significantly regulated proteins between five treatments with the
number of proteins in each segment labelled. c) Top
significantly regulated proteins with the FC difference with controls
shown. Redder boxes are more highly upregulated while bluer boxes are
more highly downregulated.
Figure 4: STRING network showing all proteins which
were significantly regulated in all treatments. Nodes represent one
STRING protein ID and edges represent a connection based on
protein-protein
interactions including known or predicted interactions based on
published literature. Solid lines connect all members of one cluster,
while dashed lines indicate connections that are not strong enough to be
included in the same cluster based on a Markov Cluster inflation factor
of 3. Central members of the Electron Transport Chain (red), glycolysis
(green), and citric acid cycle (yellow) are indicated, along with
specific proteins labelled. Insets used to more clearly display protein
names.
Figure 5: STRING networks for pejus and pessimum stress stages
and network; a) Networks with more than three nodes for all
significantly regulated proteins found in the 12-week exposure to 75g/kg
salinity representing pejus level salinity stress. Protein names in red
were upregulated, those in blue were downregulated, and those outlined
in red were upregulated in this treatment while being significantly
downregulated in extended exposures to MP at 85g/kg and/or 105g/kg.b) Networks with more than three nodes for significantly
regulated proteins found in all four pessimum level salinity stress
treatments (14-day acclimation and extended exposure to 85g/kg and
105g/kg). Legend indicates the evidence used to connect protein nodes in
each network.