Rainbow trout (Oncorhynchus mykiss) contain two calnexin genes which encode distinct proteins

Abstract

Calnexin (IP90/P88) is an integral membrane protein of the endoplasmic reticulum that binds newly synthesized N-linked glycoproteins during their folding in the ER including MHC class I molecule. This manuscript reports the identification of two unique cDNA clones of calnexin in rainbow trout. Both encode putative mature proteins of 579 and 592 aa respectively in addition to a 24 aa signal peptide. Sequence analysis revealed that only one of the two cDNA clones encodes a putative ER retention signal, K/QEDDL, followed by a serine phosphorylation site conserved with mammalian homologs. Amino acid sequence alignment illustrated conservation of the calnexin luminal domain, which consists of a globular and a P domain, in both copies. Southern blotting revealed that there are at least two copies of the calnexin gene in the trout genome and northern blotting showed a wide tissue distribution of an estimated 3 kbp calnexin transcript with an additional minor transcript of 2.3 kbp expressed only in head kidney, spleen PBLs and strongly in RTS11. Importantly, the smaller transcript was predominantly upregulated in RTS11 after a 24h treatment with the calcium ionophore A23187. In western blots, calnexin was detected primarily as a 120 kDa protein and upon A23187 treatment; a 100 kDa band was most prominently expressed. These results suggest that in salmonids there are two differentiated versions of the calnexin gene which encode proteins that may have diverged to perform unique biological functions.

Whole genome duplications have provided teleosts with many roads to peptide loaded MHC class I molecules

“…Both genes display fair expression in immunologically important tissues such as head kidney and spleen as well as other organs such as brain and ovary (Table 2 ). This gene duplication has previously been identified in trout [ 50 ] and is a remnant of the SGD displaying high sequence identity between paralogs. If one or both copies serve the classical MHCIa molecules remains to be established.…”

Transcriptomic Profiling of the Adaptive and Innate Immune Responses of Atlantic Salmon to Renibacterium salmoninarum Infection

“…As shown in Figure 13, our transcriptome profiling identified several R. salmoninarum-responsive transcripts involved in antigen-presenting and processing pathways. The teleost MH-I-dependent pathway exhibits several differences with their mammalian counterparts (123)(124)(125)(126)(127)(128); for example, different paralogues (i.e., Calnexin) and splice variants (i.e., Tapasin) of molecules involved in the MH-I pathway were identified and suggested to play diverged or unique roles in salmonid antigen presentation (123)(124)(125)(126)(127)(128). The qPCR results showed the R. salmoninarum-dependent up-regulation of fcgr1 and tnfsf14, as well as down-regulation of cdh1, flrt3, and ndrg2 (Figure 9).…”

Impacts of Low Temperature on the Teleost Immune System

“…Conversely, MHC II expression is downregulated at 2°C—but not 5°C—in rainbow trout cells, suggesting a susceptibility to bacterial diseases during cold stress [ 66 ]. Recent studies have identified, characterized and produced polyclonal antibodies to rainbow trout genes involved in the antigen processing pathway (APP) including MHC I [ 67 ], β 2 M [ 64 ], TAP1/2 [ 68 ] tapasin [ 69 ], calreticulin [ 70 , 71 ], ERp57 [ 72 ] and calnexin [ 73 ]. Collectively, these studies demonstrate the largely conserved nature of the APP in rainbow trout.…”