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Toll‐interacting protein (Tollip) is a critical regulator of TOLL‐ like receptor (TLR)‐signaling pathway. It is predominantly associated with TLR2 and TLR4 during acute inflammatory conditions and inhibits the TLR‐mediated nuclear factor‐kappa activation by suppressing the autophosphorylation of interleukin‐1 receptor‐associated kinase and its kinase activity. This article describes the Tollip of Labeo rohita (LrTollip), a highly valuable freshwater fish from the Indian subcontinent. The full‐length LrTollip complementary DNA (1412 nucleotides) encodes a 276‐amino acid (aa) protein, depicting a highly conserved target of the Myb1 (Tom1)‐binding domain (TBD; 1–53 aa), conserved core domain 2 (C2; 54–151 aa), and coupling of ubiquitin to endoplasmic reticulum degradation (CUE; 231–273 aa) domains of mouse and human counterparts. The key amino acids exerting the critical functions of Tollip, such as phospholipids recognition and ubiquitination, are present in the C2 and CUE domains of LrTollip, respectively. LrTollip is widely expressed in the kidneys, gills, spleen, liver, and blood, and among these tested tissues, the highest expression is observed in blood. In response to TLR ligands and NOD‐like receptor (NLR) ligands stimulations and Aeromonas hydrophila, Edwardsiella tarda, and Bacillus subtilis infections, LrTollip gene expression is induced in various organs/tissues with remarkable difference in their kinetics. These data together suggest the important role of LrTollip in TLR‐ and NLR‐signal transduction pathways and immune‐related diseases in fish. [ABSTRACT FROM AUTHOR]

Interleukin-1 receptor associated kinase (IRAK1) is one of the crucial signal transduction mediators in TLR/IL-1R signaling pathways in host immune system. To investigate about it in rohu (Labeo rohita), one of the economically important freshwater fish species in the Indian subcontinent, we cloned, characterized and analyzed its expression following bacterial infection and pathogens associated molecular patterns (PAMPs) stimulation. The full-length cDNA of rohu IRAK1 (LrIRAK1) consisted of 2765 nucleotide (nt) having an ORF of 2115 nt encoding a polypeptide of 704 amino acids (aa) with a molecular mass of 70.4 kDa. Structurally, LrIRAK1 consisted of twenty-nine helix, twelve strands and forty one coils making one N-terminal death domain (19–94 aa) and a central serine threonine kinase catalytic domain (or kinase domain) (188-489aa). In addition to these two prominent domains, LrIRAK1 also contained highly conserved amino acids viz., lysine 215 and aspartic acid 314 and threonine 185, 361 which were reported to be important for kinase and phosphorylation activity respectively in other animals. Similar to higher vertebrates, LrIRAK1 also consisted of CDK1 (cyclin-dependent kinase1) at 338–352 aa; NEK2 (NIMA-related kinase 2) at 47–61 aa; NEK6 (NIMA-related kinase 6) at 581–595 aa and AMPK (AMP- activated protein kinase) motif at 518–538 aa. Phylogenetically, LrIRAK1 is closely related to cave fish, common carp exhibiting high similarity (~95%) and identity (~90%). In the uninfected fish, the LrIRAK1 expression was highest in liver (~11.5 fold) and lowest in blood. In response to Aeromonas hydrophila, Edwardsiella tarda and Bacillus subtilis infection and various TLR and NLR-ligands stimulation, the expression of LrIRAK1 was markedly enhanced at various time points in almost all the tested tissues. These results together suggest the key role of LrIRAK1 in pattern recognition receptors (PRRs)-mediated host defense against pathogenic insults. • The full- length cDNA of LrIRAK1 consisted of 2765 bp, with an ORF of 2115 bp encoding 704 aa polypeptide. • LrIRAK1 protein comprised of 29 helix, 12 strands and 41 coils. • Phylogenetically LrIRAK1 was related to cave fish and common carp. • In TLR and NLR-ligands stimulation, LrIRAK1 was induced both in-vitro and in-vivo. • In A.hydrophila, E. tarda and B. subtilis infection, LrIRAK1 gene expression was significantly induced in various organs. [ABSTRACT FROM AUTHOR]