Cloning, expression, and characterization of fugu CD4, the first ectothermic animal CD4

Abstract

We have cloned and sequenced the first ectothermic animal CD4 gene from fugu, Takifugu rubripes, using a public database of the third draft sequence of the fugu genome. The fugu CD4 gene encodes a predicted protein of 463 amino acids containing four extracellular immunoglobulin (Ig)-like domains, a transmembrane region, and a cytoplasmic tail. Fugu CD4 shares low identity of about 15-20% with avian and mammalian CD4 proteins. Unlike avian and mammalian CD4, fugu CD4 lacks the Cys pair of the first Ig-like domain, but has a unique possible disulfide bond in the third domain. These differences suggest that fugu CD4 may have a different structure that could affect binding of major histocompatibility complex class II molecules and subsequent T-cell activation. In the putative fugu cytoplasmic region, the protein tyrosine kinase p56lck binding motif is conserved. The predicted fugu CD4 gene is composed of 12 exons, differing from other CD4 genes, but showing conserved synteny and many conserved sequence motifs in the promoter region. RT-PCR analysis demonstrated that the fugu CD4 gene is expressed predominantly in lymphoid tissues. We also show that fugu CD4 can be expressed on the surface of cells via transfection. Molecular characterization of CD4 in fish provides insights into the evolution of both the CD4 molecule and the immune system.

Characterization of two thymosins as immune-related genes in common carp (Cyprinus carpio L.)

“…TCR-γ, CD4 and CD8α are surface marker genes of epithelial γδT cells, Th and Tc cells. Expression of CD4 and CD8 genes are essential for cell-mediated immune response and the development of T lymphocytes (Suetake et al, 2004). Up-regulation of these lymphocytes-related gene expression indicated that overexpression of carp ProTα b and Tβ-l may induce the development of T lymphocytes, and ultimately enhance the cell mediated immune response.…”

Cloning, characterization, and expression pattern of Atlantic halibut (Hippoglossus hippoglossus L.) CD4-2, Lck, and ZAP-70

“…The genomic organization of the halibut CD4-2 gene shows a close relationship with CD4 and LAG-3 genes, as the first domain D1 was shown to be dispersed between two exons. This is a special feature seen in the CD4 and LAG-3 genes of higher vertebrates, and found in all the teleost CD4 genes characterized to date [3][4][5][6][7][8].…”

“…Surprisingly, the F 43 was conserved in halibut CD4-2 ( Figure 2), and not in the classical four-domain halibut CD4 [3]. The two-domain CD4 of fugu is also in the possession of a phenylalanine corresponding to human F 43 [4], giving the consensus FxxK in halibut, fugu, and human but not in other CD4 like genes [3][4][5][6][7][8][9][10]. Other features common in mammalian and fish CD4 Igdomains are the presence of cysteines for intra-chain disulfide bridging and a core tryptophan forming an essential structural triad thought to stabilize most Ig-domains [35].…”

“…Molecules identified in halibut believed to have preserved their structural characteristics includes RAG1 (FJ769824); TCRα, TCRβ, CD3γδ, CD3ε, CD3ζ [1]; CD8α, CD8β [2]; and CD4 [3]. In fish, different CD4 like molecules have been identified that shows resemblance to CD4 of higher vertebrates [4][5][6][7][8][9][10], one form that has the classical four Ig-like domain (D1-D4) structure, another that has three Ig-domains (D1-D3), and a third form that holds only two Ig-domains (D1-D2) followed by a connecting peptide that is believed to be compensating for the two missing Ig-domains. Seemingly, different fish species are in the possession of only two of the three divergent forms of CD4, the classical CD4 and either the three domain form or the two domain form.…”

Teleost B7 Expressed on Monocytes Regulates T Cell Responses

“…Dendritic cells (DCs), 4 monocytes/macrophages, and B cells have been identified as major professional APCs (1,2). These APCs capture and process Ags and then present the processed antigenic peptides upon MHC molecules to the T cells.…”