Genes differentially expressed in response to salinity
(comparison 1 and 2)
When contrasting SwC and SwFw (comparison 1), a total of 10 transcripts
were significantly differentially regulated, six which were annotated.
Of the 10 transcripts, 3 transcripts were significantly different only
for comparison 1 (Figure 4 ). Two of the transcripts were
annotated; a gene predicted to be involved in the elongation factor
process (si:ch211-13k12.2 ) and the enzyme Galactosylceramide
sulfotransferase (GAL3ST1 ). Five transcripts were shared with
comparison 2: an arrestin domain (arrdc3a ), a solute carrier
(SLC16a9a) (Figure 6a ), a cytochrome P450, family 1
(CYP1a ) (Figure 6b ), a docking protein (dok4 ) and
a novel gene, ENSGACG00000019379, which shares sequence homology
with TBC1 family member 24 (BlastP GenBank) that has a potential
function in intracellular trafficking. Two additional transcripts, which
were also not annotated, were shared with comparison 3 (Figure
4 ).
Of the 1535 genes reported in comparison 2, between freshwater ecotypes,
a total of 691 transcripts (569 annotated) were upregulated in FwSw when
compared to FwC (Table 1 ). The upregulated gene-list included
several ion transporting ATPases; ATPase Na+/K+ transporting subunit
beta 1a (ATP1b1a ), ATPase sarcoplasmic/endoplasmic reticulum Ca2+
transporting 1 (ATP2a1 , Figure 6b ) and ATPase
phospholipid transporting 11C (ATP11c ); t wo
ABC-transporter genes were upregulated in FwSw (ABCf2a andABCb6a ) which belong to a family of proteins that utilize the
energy of ATP binding and hydrolysis to transport various substrates
across cellular membranes. A total of 20 different solute carriers were
also upregulated in FwSw, and two genes involved in vesicular
trafficking; transmembrane emp24 trafficking protein 2 (tmed2 ,Figure 6b ) and 10 (tmed10 ). Genes linked to stress
included five Heat Shock Protein (HSP)-genes (HSPa4b, HSPa5,
HSPa8 (Figure 6b ), HSPa8b, HSPd1 ) and seven ubiquitin
specific peptidase (USP)-genes. The HSPa8 and HSPa8bencodes members of HSP70, and four DnaJ homologs which are
co-chaperones of the HSP70-family were also upregulated in FwSw
(dnaja1, dnajb1a, dnaja2a, dnajb9b ). Gene ontology enrichment of
the upregulated genes involved processes such as peptide biosynthetic
process, cellular response to xenobiotic stimulus and glutathione
metabolic process (Table 2 ).
A total of 671 annotated genes were downregulated in FwSw when compared
to FwC (Table 1 ). Several known genes had reduced expression
for freshwater fish when in saltwater; six genes related to ion
transporting ATPases, including five related to phospholipid
transporting (ATP8a1, ATP8b2, ATP9a, ATP9b and ATP10d ),
and one linked to metallopeptidase and ATP-synthase assembly factor
homologs (ATP23 ). Further, seven different solute carriers were
downregulated, as were genes linked to the potassium voltage-gated
channel (KCNd3 ), the potassium inwardly rectifying channel
(KCNj2a ), a potassium channel tetramerisation domain
(KCTd12b ) and the chloride channel, voltage-sensitive 6
(CKCn6 ). Two genes linked to calcium regulation; an EF-hand
calcium binding domain (EFcab7 ) and a transient receptor
potential cation channel gene (TRMP7 ), and two genes linked to
thyroid hormone signalling, thyroid hormone receptor interactor 10a
(trip10a ) and thyroid hormone receptor interactor 4
(trip4 ). Related to osmosensing, and involved in the activation
and differentiation of immune cells, five interlukins all had similarly
expression across groups, except for FwSw where they were downregulated
(ILf3a, ILf3b, IL113ra2, IL15, and IL19 ). No HSP-genes were
downregulated, but one ATP-binding cassette (ABCh1 ), three DnaJ
homologs (co-chaperones of the HSP70- family: dnajc7, dnajc9 anddnajc11b ), together with three USP-genes (USP31, USP40 and
USP45 ) all had lower expression in FwSw. When testing all downregulated
genes in this comparison, transcripts for GO-enrichments processes such
as DNA repair, nucleosome assembly, cilium assembly were included on top
of the list (Table 2 ).