The CD100 Receptor Interacts with Its Plexin B2 Ligand to Regulate Epidermal γδ T Cell Function

Summaryγδ T cells respond rapidly to keratinocyte damage, providing essential contributions to the skin wound healing process. The molecular interactions regulating their response are unknown. Here, we identify a role for interaction of plexin B2 with the CD100 receptor in epithelial repair. In vitro blocking of plexin B2 or CD100 inhibited γδ T cell activation. Furthermore, CD100 deficiency in vivo resulted in delayed repair of cutaneous wounds due to a disrupted γδ T cell response to keratinocyte damage. Ligation of CD100 in γδ T cells induced cellular rounding via signals through ERK kinase and cofilin. Defects in this rounding process were evident in the absence of CD100-mediated signals, thereby providing a mechanistic explanation for the defective wound healing in CD100-deficient animals. The discovery of immune functions for plexin B2 and CD100 provides insight into the complex cell-cell interactions between epithelial resident γδ T cells and the neighboring cells they support

A High Density SNP Array for the Domestic Horse and Extant Perissodactyla: Utility for Association Mapping, Genetic Diversity, and Phylogeny Studies

An equine SNP genotyping array was developed and evaluated on a panel of samples representing 14 domestic horse breeds and 18 evolutionarily related species. More than 54,000 polymorphic SNPs provided an average inter-SNP spacing of ∼43 kb. The mean minor allele frequency across domestic horse breeds was 0.23, and the number of polymorphic SNPs within breeds ranged from 43,287 to 52,085. Genome-wide linkage disequilibrium (LD) in most breeds declined rapidly over the first 50–100 kb and reached background levels within 1–2 Mb. The extent of LD and the level of inbreeding were highest in the Thoroughbred and lowest in the Mongolian and Quarter Horse. Multidimensional scaling (MDS) analyses demonstrated the tight grouping of individuals within most breeds, close proximity of related breeds, and less tight grouping in admixed breeds. The close relationship between the Przewalski's Horse and the domestic horse was demonstrated by pair-wise genetic distance and MDS. Genotyping of other Perissodactyla (zebras, asses, tapirs, and rhinoceros) was variably successful, with call rates and the number of polymorphic loci varying across taxa. Parsimony analysis placed the modern horse as sister taxa to Equus przewalski. The utility of the SNP array in genome-wide association was confirmed by mapping the known recessive chestnut coat color locus (MC1R) and defining a conserved haplotype of ∼750 kb across all breeds. These results demonstrate the high quality of this SNP genotyping resource, its usefulness in diverse genome analyses of the horse, and potential use in related species

cDNA Sequence and Fab Crystal Structure of HL4E10, a Hamster IgG Lambda Light Chain Antibody Stimulatory for γδ T Cells

Hamsters are widely used to generate monoclonal antibodies against mouse, rat, and human antigens, but sequence and structural information for hamster immunoglobulins is sparse. To our knowledge, only three hamster IgG sequences have been published, all of which use kappa light chains, and no three-dimensional structure of a hamster antibody has been reported. We generated antibody HL4E10 as a probe to identify novel costimulatory molecules on the surface of γδ T cells which lack the traditional αβ T cell co-receptors CD4, CD8, and the costimulatory molecule CD28. HL4E10 binding to γδ T cell, surface-expressed, Junctional Adhesion Molecule-Like (JAML) protein leads to potent costimulation via activation of MAP kinase pathways and cytokine production, resulting in cell proliferation. The cDNA sequence of HL4E10 is the first example of a hamster lambda light chain and only the second known complete hamster heavy chain sequence. The crystal structure of the HL4E10 Fab at 2.95 Å resolution reveals a rigid combining site with pockets faceted by solvent-exposed tyrosine residues, which are structurally optimized for JAML binding. The characterization of HL4E10 thus comprises a valuable addition to the spartan database of hamster immunoglobulin genes and structures. As the HL4E10 antibody is uniquely costimulatory for γδ T cells, humanized versions thereof may be of clinical relevance in treating γδ T cell dysfunction-associated diseases, such as chronic non-healing wounds and cancer

Genome-Wide Analysis Reveals Selection for Important Traits in Domestic Horse Breeds

Peer reviewe

Structure prediction and automated modeling of HL4E10.

<p>Cartoon representation of the superimposition of the experimentally determined HL4E10 heavy and light chain variable domain structures and the three top scoring heavy (1W72, 2G75, 1ADQ) and light chain (1A7P, 1GIG, and 1DL7) computational models. Heavy and light chains are shown in dark and light gray, respectively. The CDR loops of HL4E10 are color coded as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone-0019828-g001" target="_blank">Fig. 1</a>&<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone-0019828-g002" target="_blank">2</a>: CDR L1 yellow, CDR L2 cyan, CDR L3 orange, CDR H1 blue, CDR H2 pink, CDR H3 green. The Cα atoms of the experimentally determined HL4E10 structure and the computational models superimpose well, with an average r.m.s.d. of 0.68 Å for the heavy chains and 0.77 Å for the light chains, respectively. The largest deviations are observed, as expected, in the CDR loops, namely L1, L3 and H3.</p

Amino-acid sequence alignment of the HL4E10 hamster IgG heavy chain with those of other hamster antibodies.

<p>The HL4E10 heavy chain is aligned with H28.710 (U17166 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone.0019828-Collins1" target="_blank">[33]</a>), 145.2c11 (U17871 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone.0019828-Alegre1" target="_blank">[31]</a>), and 3A5-1 (S80616 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone.0019828-Mallender1" target="_blank">[30]</a>). Color coding and shading is used as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone-0019828-g001" target="_blank">Fig. 1</a>, CDR H1 is in blue, CDR H2 in pink, CDR H3 in green, and the glycine-, proline-, cysteine-rich hinge region between V_HC_H1 and C_H2C_H3 is shaded gray.</p

Hierarchical clustering of the HL4E10 protein sequence with known immunoglobulins.

<p>Distances were calculated between protein sequences of (<b>A</b>) heavy chains and (<b>B</b>) light chains as the number of amino acid substitutions per site. Scale bar indicates distances. GenBank accession numbers of sequences included in the analysis are indicated within branch labels. Hierarchical clustering was performed on distance matrices generated from protein sequences with removed signal peptides.</p

Amino-acid sequence alignment of the HL4E10 hamster IgG light chain with those of other hamster antibodies.

<p>The HL4E10 lambda light chain is aligned with H28.710 (Genbank accession no. U17165 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone.0019828-Whitters1" target="_blank">[32]</a>), 145.2c11 (U17870 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone.0019828-Alegre1" target="_blank">[31]</a>), and 1F4 (S80615 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019828#pone.0019828-Mallender1" target="_blank">[30]</a>). Identical residues are in red and homologous exchanges are in green. Signal peptides are underlined, CDR loops are shaded: CDR L1 is in yellow, CDR L2 is in cyan, CDR L3 is in orange-red, and residues which are rarely observed in antibodies at particular locations are shaded gray. Kabat numbering is used throughout, as well as the definition of CDRs.</p