**2. Osmoregulatory genes**

#### **2.1. Na+ /K+ -ATPase**

**Figure 3.** Differentially expressed genes from forward SSH library of gill tissues of *P. monodon* under low (3 ppt) and

**Figure 4.** Differentially expressed genes from the reverse SSH library of gill tissues of *P. monodon* under low (3 ppt) and

high (55 ppt) salinity conditions

104 Abiotic and Biotic Stress in Plants - Recent Advances and Future Perspectives

high (55 ppt) salinity conditions

Na+ /K+ -ATPase, a transmembrane protein, contains three subunits, α-, β, and γ- subunit, which are involved in exporting three Na<sup>+</sup> from cytosol in exchange for two K<sup>+</sup> or NH4 + from extrac‐ ellular fluid for each ATP hydrolyzed. The crustacean α-subunit, which is 71–74% identical in amino acid sequence to those of vertebrate α-subunit sequences, binds to ATP and functions for the catalytic action of the enzyme [12]. The binding of Na<sup>+</sup> /K+ -ATPase complex to basolat‐ eral membrane requires participation of β-subunit and the γ-subunits of enzyme [13]. The activity of Na+ +K+ -ATPase in gill tisues of crustaceans depends on the osmoconcentration gradient occurring between hemolymph and the external medium. In crustaceans, there is an increase in Na+ /K+ -ATPase activity when transferred from natural seawater to dilute seawater [14-15]. With the lowering of salinity when compared to that of normal seawater, the euryha‐ line crustacea undergo hyperosmoregulation. Increased enzymatic activity of Na<sup>+</sup> +K+ -ATPase and increased α-subunit gene expression has been observed in the gill tissues of crabs [12]. Substantial increase in Na+ /K+ -ATPase specific activity (300%), Na+ /K+ -ATPase protein levels (200%), and gene expression level of α-subunit (150%) has been observed in blue crab *Callinectes sapidus* crabs during acclimitization to dilute seawater of 10 ppt salinity [16]. *P. monodon* reared in 7 ppt seawater showed drastic morphological alterations of the antennal glands. The shrimps also showed higher expression and activity of the enzyme Na<sup>+</sup> /K+ -ATPase in the antennal glands under low salinity conditions [17]. *L. vannamei* when transferred to different low salinity conditions ranging from 15 ppt to 1 ppt revealed no significant difference within 3 h for Na+ /K+ -ATPase α-subunit gene expression and enzyme activity. However, there was a rapid increase at 6 h followed by decrease in the expression level from 12 h to 24 h suggesting Na<sup>+</sup> /K+ -ATPase is stimulated by salinity stress [18]. The study involving V-H ATPase α -subunit and Na<sup>+</sup> /K+ -ATPase β-subunit response to environmental stress (bacteria, pH, Cd, salinity, and low temperature) revealed both the genes to be responsive to these environmental stress conditions. However, the V-H ATPase α -subunit and the Na<sup>+</sup> /K+ -ATPase β-subunit, which is involved in proper folding and transport of Na<sup>+</sup> /K+ -ATPase enzyme, were found to be more sensitive to salinity stress when compared to other stress factors. The exposure of *L. vannamei* to salinity stress resulted in significant changes in the expression of V-H ATPase α-subunit and Na<sup>+</sup> /K+ -ATPase β-subunit gene expression levels in the hepato‐ pancreas and gills of the shrimp. Na<sup>+</sup> /K+ -ATPase β-subunit gene expression after exposure to 5 ppt increased to a highest level (17-fold) at 12 h in the gill tissues, whereas, in hepatopancreas the maximum gene expression levels (4.4-fold) were observed 6 h after exposure to 10 ppt salinity conditions [19].

At low (3 ppt) salinity conditions stress conditions, significant increase in the Na<sup>+</sup> /K+ -ATPase α-subunit gene expression levels was observed in gill (34.28-fold) tissues of *P. monodon* [20]. At higher salinity stress of 55 ppt, *P. monodon* Na+ /K+ -ATPase gene responded to salinity stress conditions with significant expression levels in gill (15.23-fold) tissues [21]. These results suggests that *P. monodon* Na+ /K+ -ATPase gene is involved in osmoregulatory process in shrimp and responds significantly under salinity stress.
