Molecular Characterization of MHC Class I Alpha 1 and 2 Domains in Asian Seabass (Lates calcarifer) - PubMed
- ️Sat Jan 01 2022
Molecular Characterization of MHC Class I Alpha 1 and 2 Domains in Asian Seabass (Lates calcarifer)
Zhixuan Loh et al. Int J Mol Sci. 2022.
Abstract
The Asian seabass is of importance both as a farmed and wild animal. With the emergence of infectious diseases, there is a need to understand and characterize the immune system. In humans, the highly polymorphic MHC class I (MHC-I) molecules play an important role in antigen presentation for the adaptive immune system. In the present study, we characterized a single MHC-I gene in Asian seabass (Lates calcarifer) by amplifying and sequencing the MHC-I alpha 1 and alpha 2 domains, followed by multi-sequence alignment analyses. The results indicated that the Asian seabass MHC-I α1 and α2 domain sequences showed an overall similarity within Asian seabass and retained the majority of the conserved binding residues of human leukocyte antigen-A2 (HLA-A2). Phylogenetic tree analysis revealed that the sequences belonged to the U lineage. Mapping the conserved binding residue positions on human HLA-A2 and grass carp crystal structure showed a high degree of similarity. In conclusion, the availability of MHC-I α1 and α2 sequences enhances the quality of MHC class I genetic information in Asian seabass, providing new tools to analyze fish immune responses to pathogen infections, and will be applicable in the study of the phylogeny and the evolution of antigen-specific receptors.
Keywords: Asian seabass; Lates calcarifer; MHC class I; polymorphism; sequence variability.
Conflict of interest statement
The authors declare no conflict of interest.
Figures

Alignment of Asian seabass MHC-I α1 and α2 amino acid sequences with other representative MHC-I sequences. Multiple sequence alignment of Asian amino acid sequences with other representative fish MHC-I and human HLA sequences. Black indicates similar residues that were probably found before the molecular evolution of MHC classes I and II, blue indicates similar residues that are common in classical MHC-I, and red indicates conserved binding residues in classical MHC-I. The sequences are divided into α1 and α2 regions, which correspond to exon 2 and exon 3, respectively. The residue positions indicated below the alignment are based on the HLA-A2 protein. Structural indications S denotes the β-strand and H denotes the helix based on the pHLA-A2 structure (PDB database accession 3PWN). The sequence references are as follows: Orni (Oreochromis niloticus, Nile tilapia) -HAA (TSA: GBAZ01123113); Gaac (Gasterosteus aculeatus, Stickleback) -HAA (DW655318); Lacr (Larimichthys crocea, Yellow croaker) –HAA (XP_010741942.1); Pore (Poecilia reticulata, Guppy) -HAA (XP_008432358.1 and TSA: GFHH01045885); Teni (Tetraodon nigroviridis, Tetraodon) -HAA (CAG07665.1); Sasa (Salmo salar, Atlantic salmon) -SAA (ACY30362.1), -ZAAa (ACX35596.1), and -UBA (*0301, XP_014032819); Leoc (Lepisosteus oculatus, Spotted gar) -LO14 (TSA: GFIM01040660), -LO12 (JH591577:52,184-56,541), and -U (TSA: GFIM01032149); Ctid (Ctenopharyngodon idella, Grass carp) -UAA (5H5Z-A); Asme (Astyanax mexicanus, Mexican tetra) -AM33 (ENSAMXG00000017444) and -P (TSA: GFIF01000014); Taru (Takifugu rubripes, Fugu) -TR16 (Scaffold_497:44,782-49,340); HLA (Human leukocyte antigen) -A2 (AAA76608.2); -B15:01 (HG794370.1), -C1:02 (HG794388.1), and Lca17733 (Lates calcarifer, Asian seabass) (XM_018684417). SG denotes samples obtained from Singapore and AU denotes samples obtained from Australia.

Amino acid identity and sequence variability within the Asian seabass α1 and α2 domains. (a) Heatmap of amino acid similarity values for Asian seabass and Lca17733 MHC-I α1 and α2 domains. (b) Sequence variability calculated using the variability metric (V) analysis based on the Shannon entropy equation. Sites with V >1 are considered as highly polymorphic in the sequence alignment of the Asian seabass and Lca17733 MHC-I α1 and α2 domains.

Phylogenetic trees based on α1 and α2 domain amino acid sequences of Asian seabass and other representative teleost fishes’ MHC-I molecules. Neighbor-joining tree [32] of α1 (left tree) and α2 (right tree) domain sequences. The trees are drawn to scale, with branch lengths indicating the number of amino acid substitutions per site. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) [33] is shown on each node. The evolutionary distances were computed using the Poisson correction method [34] and are presented in the unit of the number of amino acid substitutions per site. The analysis involved 32 amino acid sequences. There were 99 positions in each of the final datasets. Evolutionary analyses were conducted in MEGA11 [35]. Clusters with the six different MHC class I lineages found in teleosts are shown with colored boxes. The sequence references are listed in Figure 1.

Schematic crystal structure of the human HLA-A*02:01 (PDB code 3MRE) and Ctid-UAA (PDB code 5H5Z) peptide binding groove defined by the α1 and α 2 domains. HLA-A*02:01 structure (a,b, pink) and Ctid-UAA structure (c,d, grey) with similar residues from the sequence of representative Asian seabass MHC-I residues mapped viewed from the top (a,c) and the PΩ side (b,d). Black indicates similar residues that were probably found before the molecular evolution of MHC classes I and II, blue indicates similar residues that are common in classical MHC-I, and red indicates conserved binding residues in classical MHC-I.
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