Biomedical subjects

Kerry J Laing

Publications and source records attributed to Kerry J Laing.

6 recordsLinked to original sources

PubMed2006

Molecular and biochemical analysis of rainbow trout LCK suggests a conserved mechanism for T-cell signaling in gnathostomes.

Two genes were identified in rainbow trout that display high sequence identity to vertebrate Lck. Both of the trout Lck transcripts are associated with lymphoid tissues and were found to be highly expressed in IgM-negative lymphocytes. In vitro analysis of trout lymphocytes indicates that trout Lck mRNA is up-regulated by T-cell mitogens, supporting an evolutionarily conserved function for Lck in the signaling pathways of T-lymphocytes. Here, we describe the generation and characterization of a specific monoclonal antibody raised against the N-terminal domains of recombinant trout Lck that can recognize Lck protein(s) from trout thymocyte lysates that are similar in size ( approximately 57kDa) to mammalian Lck. This antibody also reacted with permeabilized lymphocytes during FACS analysis, indicating its potential usage for cellular analyses of trout lymphocytes, thus representing an important tool for investigations of salmonid T-cell function.

Amino Acid Sequence↗

PubMed2006

Evolution of the CD4 family: teleost fish possess two divergent forms of CD4 in addition to lymphocyte activation gene-3.

The T cell coreceptor CD4 is a transmembrane glycoprotein belonging to the Ig superfamily and is essential for cell-mediated immunity. Two different genes were identified in rainbow trout that resemble mammalian CD4. One (trout CD4) encodes four extracellular Ig domains reminiscent of mammalian CD4, whereas the other (CD4REL) codes for two Ig domains. Structural motifs within the amino acid sequences suggest that the two Ig domains of CD4REL duplicated to generate the four-domain molecule of CD4 and the related gene, lymphocyte activation gene-3. Here we present evidence that both of these molecules in trout are homologous to mammalian CD4 and that teleosts encode an additional CD4 family member, lymphocyte activation gene-3, which is a marker for activated T cells. The syntenic relationships of similar genes in other teleost and non-fish genomes provide evidence for the likely evolution of CD4-related molecules in vertebrates, with CD4REL likely representing the primordial form in fish. Expression of both CD4 genes is highest in the thymus and spleen, and mRNA expression of these genes is limited to surface IgM- lymphocytes. consistent with a role for T cell functionality. Finally, the intracellular regions of both CD4 and CD4REL possess the canonical CXC motif involved in the interaction of CD4 with p56LCK, implying that similar mechanisms for CD4+ T cell activation are present in all vertebrates. Our results therefore raise new questions about T cell development and functionality in lower vertebrates that cannot be answered by current mammalian models and, thus, is of fundamental importance for understanding the evolution of cell-mediated immunity in gnathosomes.

Amino Acid Sequence↗

PubMed2004

Chemokines.

Chemokines are small proteins that control cellular migration. An extensive family of these molecules has been described in mammals containing nearly 50 members. Within this family are four groups, each defined by the different spacing of two N-terminal cysteines, which form disulphide bonds with two other cysteine residues to create the tertiary structure characteristic of chemokines. Recent evidence shows the chemokine family is not unique to mammals, with several members also identified in birds, amphibians and fish, including a primitive vertebrate, the lamprey. Although there is less evidence to define the roles of chemokines in these lower vertebrates, structural similarities allow some predictions to their function, against which further studies are being made. Additionally, some microorganisms (particularly viruses) appear to have copied genes for chemokines, presumably to confuse the immune system of their host. This review aims to bring together the current information concerning identified chemokines throughout vertebrates and microorganisms.

Amino Acid Sequence↗

PubMed2004

Trout CC chemokines: comparison of their sequences and expression patterns.

Several thousand EST sequences were recently made available in the EMBL sequence database from the rainbow trout Oncorhynchus mykiss. BLAST based searches were utilised to identify sequences resembling mammalian CC chemokines within these ESTs. Fifteen new and unique CC chemokine-like sequences were identified for trout, bringing the total of known CC chemokine sequences in trout to 18 when including those already published. Some of these trout chemokines appeared highly related (in pairs) suggesting recent duplication events or tight evolutionary constraints. Phylogenetically, the trout chemokine sequences grouped with both inducible and constitutive mammalian CC chemokine subtypes, suggesting early divergence of these functional groups. Expression analyses on gill and head kidney show constitutive expression of many of these trout CC chemokines in these lymphoid-rich tissues. However, induction of some of the chemokines structurally related to 'inducible' CC chemokines was observed in a trout macrophage-like cell line (RTS-11) in response to stimulation with recombinant TNFalpha.

Amino Acid Sequence↗

PubMed2002

Identification and analysis of an interleukin 8-like molecule in rainbow trout Oncorhynchus mykiss.

An interleukin 8 (IL-8) homologue has been identified in the rainbow trout Oncorhynchus mykiss. The transcript contains an open reading frame of 294 nucleotides that translates into a 97 amino acid putative peptide, with 5' and 3' untranslated regions (UTR) of 171 and 453 nucleotides, respectively. As with previously sequenced lamprey and flounder genes, the trout amino acid sequence lacks the typical ELR motif upstream of the first pair of cysteines, where DLR is present. The trout IL-8 gene contains four exons divided by three short introns of 341, 247 and 292bp, and occupies 1824bp of genomic DNA. RT-PCR reveals a low level constitutive expression of the IL-8 homologue in many tissues, including spleen, heart, liver, head kidney and gill. Expression was not detectable in the brain. Whilst no apparent affect of lipopolysaccharide (LPS) on IL-8 expression was observed in vivo, stimulation of a trout macrophage cell line (RTS-11) with either LPS or poly I:C did result in clear up-regulation of IL-8 expression, detectable by RT-PCR and Northern blot analysis.

Amino Acid Sequence↗

PubMedDate not supplied

A CXC chemokine sequence isolated from the rainbow trout Oncorhynchus mykiss resembles the closely related interferon-gamma-inducible chemokines CXCL9, CXCL10 and CXCL11.

A sequence encoding a CXC - type chemokine from rainbow trout was found to most resemble members of the CXCL9/CXCL10/CXCL11 sub-family. In mammals, all 3 chemokines are regulated by IFN-gamma and are chemotactic for activated T lymphocytes. The trout chemokine (gammaIP1), with a message of 787 nucleotides, contains 100 amino acids in a typical non-ELR CXC chemokine arrangement. A second sequence (gammaIP2), with 6 nucleotide differences in the coding region when compared to the first, was also identified although it is not known whether this is a second functional gene or a second allele. The gene is separated onto 4 exons, and the introns intervene in conserved positions according to the mammalian equivalents. The sequence encoded by the second exon shares the highest amino acid identity (37%) with CXCL10, with lower values of identity to other CXC chemokines (17-31%). Furthermore, phylogenetic analysis groups the trout chemokine with mammalian CXCL9, CXCL10 and CXCL11 peptides. Constitutive expression of gammaIP is seen in trout gill and low level expression in spleen, head kidney and liver. In RTS-11 cells, gammaIP expression can be induced with poly I:C, but not by LPS, suggesting virus-mediated regulation of gammaIP. Intraperitoneal injection of recombinant trout TNF-alpha caused elevation in gammaIP mRNA levels in trout head kidney.

Amino Acid Sequence↗