TAPASIN, a V-C1 (variable-constant) immunoglobulin superfamily (IgSF) molecule that links MHC class I molecules to the transporter associated with antigen processing (TAP) in the endoplasmic reticulum (ER) is encoded by the TAPBP gene, located near to the MHC at 6p21.3. A related gene was identified at chromosome position 12p13.3 between the CD27 and VAMP1 genes near a group of MHC-paralogous loci. The gene, which we have called TAPBP-R (R for related), also encodes a member of the IgSF, TAPASIN-R. Its putative product contains similar structural motifs to TAPASIN, with some marked differences, especially in the V domain, transmembrane and cytoplasmic regions. By using the mouse ortholog to screen tissue, we revealed that the TAPBP-R gene was broadly expressed. Sub-cellular localization showed that the bulk of TAPASIN-R is located within the ER but biotinylation experiments were consistent with some expression at the cell surface. TAPASIN-R lacks an obvious ER retention signal. The function of TAPASIN-R will be of interest in regards to the evolution of the immune system as well as antigen processing.

No abstract

G protein-coupled receptors (GPCRs) have a key role in many biological processes and are important drug targets for many human diseases. Therefore, understanding the molecular interactions between GPCRs and their ligands would improve drug design. Here, we describe an approach that allows the rapid identification of functional agonists expressed in bacteria. Transgenic Caenorhabditis elegans expressing the human chemokine receptor 5 (CCR5) in nociceptive neurons show avoidance behavior on encounter with the ligand MIP-1␣ and avoid feeding on Escherichia coli expressing MIP-1␣ compared with control bacteria. This system allows a simple activity screen, based on the distribution of transgenic worms in a binary food-choice assay, without a requirement for protein purification or tagging. By using this approach, a library of 68 MIP-1␣ variants was screened, and 13 critical agonist residues involved in CCR5 activation were identified, four of which (T8, A9, N22, and A25) have not been described previously, to our knowledge. Identified residues were subsequently validated in receptor binding assays and by calcium flux assays in mammalian cells. This approach serves not only for structure/function studies as demonstrated, but may be used to facilitate the discovery of agonists within bacterial libraries. CCR5 ͉ MIP-1␣G PCRs constitute the largest family of mammalian cell surface proteins. They have central roles in physiological processes and represent major targets for current pharmaceutical therapies. GPCRs bind a diverse set of ligands including volatile odorants, hormones, and protein ligands such as chemokines and cytokines, which represent a major medically relevant class of GPCR ligands. Caenorhabditis elegans is a bacteria-feeding soil nematode whose strategies for detecting food sources and environmental compounds include using GPCRs expressed in gustatory neurons driving ''hard-wired'' repulsive or attractive responses. Thus, C. elegans provides a potential means of studying GPCR activation. We have previously shown that heterologous expression of GPCRs in this system permit changes in specificity of the avoidance response. Heterologous expression of Somatostatin Receptor 2 and CCR5 in the chemosensory nociceptive neurons ASH and ADL of C. elegans resulted in avoidance behavior when the transgenic worms were exposed to somatostatin and MIP-1␣, respectively (1).The work described above used purified soluble compounds placed in the path of the worm to elicit an avoidance response. Because C. elegans feeds on bacteria including Escherichia coli, and because gustatory neurons have an important role in taste perception and food preference, we reasoned that expression of functional MIP-1␣ in bacteria (2) could drive an avoidance response in the transgenic animals, leading to reduced feeding on bacteria expressing the agonist. Here, we demonstrate that CCR5 transgenic C. elegans exhibit avoidance feeding behavior toward bacteria expressing functional MIP-1␣ compared with bacteria expressing an unrelated protein in a...

Background: G-protein-coupled receptors (GPCRs) play a crucial role in many biological processes and represent a major class of drug targets. However, purification of GPCRs for biochemical study is difficult and current methods of studying receptor-ligand interactions involve in vitro systems. Caenorhabditis elegans is a soil-dwelling, bacteria-feeding nematode that uses GPCRs expressed in chemosensory neurons to detect bacteria and environmental compounds, making this an ideal system for studying in vivo GPCR-ligand interactions. We sought to test this by functionally expressing two medically important mammalian GPCRs, somatostatin receptor 2 (Sstr2) and chemokine receptor 5 (CCR5) in the gustatory neurons of C. elegans.

While the majority of children infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) display mild or no symptoms, rare individuals develop severe disease presenting with multisystem inflammatory syndrome (MIS-C). The reason for variable clinical manifestations is not understood. Here, we carried out TCR sequencing and conducted comparative analyses of TCR repertoires between children with MIS-C (n = 12) and mild (n = 8) COVID-19. We compared these repertoires with unexposed individuals (samples collected pre-COVID-19 pandemic: n = 8) and with the Adaptive Biotechnologies MIRA dataset, which includes over 135,000 high-confidence SARS-CoV-2-specific TCRs. We show that the repertoires of children with MIS-C are characterised by the expansion of TRBV11-2 chains with high junctional and CDR3 diversity. Moreover, the CDR3 sequences of TRBV11-2 clones shift away from SARS-CoV-2 specific T cell clones, resulting in distorted TCR repertoires. In conclusion, our study reports that CDR3-independent expansion of TRBV11-2+ cells, lacking SARS-CoV-2 specificity, defines MIS-C in children.