Identification of Class I HLA T Cell Control Epitopes for West Nile Virus

The recent West Nile virus (WNV) outbreak in the United States underscores the importance of understanding human immune responses to this pathogen. Via the presentation of viral peptide ligands at the cell surface, class I HLA mediate the T cell recognition and killing of WNV infected cells. At this time, there are two key unknowns in regards to understanding protective T cell immunity: 1) the number of viral ligands presented by the HLA of infected cells, and 2) the distribution of T cell responses to these available HLA/viral complexes. Here, comparative mass spectroscopy was applied to determine the number of WNV peptides presented by the HLA-A*11:01 of infected cells after which T cell responses to these HLA/WNV complexes were assessed. Six viral peptides derived from capsid, NS3, NS4b, and NS5 were presented. When T cells from infected individuals were tested for reactivity to these six viral ligands, polyfunctional T cells were focused on the GTL9 WNV capsid peptide, ligands from NS3, NS4b, and NS5 were less immunogenic, and two ligands were largely inert, demonstrating that class I HLA reduce the WNV polyprotein to a handful of immune targets and that polyfunctional T cells recognize infections by zeroing in on particular HLA/WNV epitopes. Such dominant HLA/peptide epitopes are poised to drive the development of WNV vaccines that elicit protective T cells as well as providing key antigens for immunoassays that establish correlates of viral immunity. © 2013 Kaabinejadian et al

Determination of Cellular Lipids Bound to Human CD1d Molecules

CD1 molecules are glycoproteins that present lipid antigens at the cell surface for immunological recognition by specialized populations of T lymphocytes. Prior experimental data suggest a wide variety of lipid species can bind to CD1 molecules, but little is known about the characteristics of cellular ligands that are selected for presentation. Here we have molecularly characterized lipids bound to the human CD1d isoform. Ligands were eluted from secreted CD1d molecules and separated by normal phase HPLC, then characterized by mass spectroscopy. A total of 177 lipid species were molecularly identified, comprising glycerophospholipids and sphingolipids. The glycerophospholipids included common diacylglycerol species, reduced forms known as plasmalogens, lyso-phospholipids (monoacyl species), and cardiolipins (tetraacyl species). The sphingolipids included sphingomyelins and glycosylated forms, such as the ganglioside GM3. These results demonstrate that human CD1d molecules bind a surprising diversity of lipid structures within the secretory pathway, including compounds that have been reported to play roles in cancer, autoimmune diseases, lipid signaling, and cell death

Size dependence of the pressure-induced γ to α structural phase transition in iron oxide nanocrystals

The size trend for the pressure-induced γ-Fe₂O₃ (maghemite) to α-Fe₂O₃ (haematite) structural phase transition in nanocrystals has been observed. The transition pressure was found to increase with decreasing nanocrystal size: 7 nm nanocrystals transformed at 27 ± 2 GPa, 5 nm ones at 34 ± 3 GPa and 3 nm ones at 37 ± 2 GPa. Annealing of a bulk sample of γ-Fe₂O₃ was found to reduce the transition pressure from 35 ± 2 to 24 ± 2 GPa. The bulk modulus was determined as 262 ± 6 GPa for 7 nm nanocrystals of γ-Fe₂O₃, which is significantly higher than the value of 190 ± 6 GPa that we measured for bulk samples. For α-Fe₂O₃, the bulk moduli for 7 nm nanocrystals (336 ± 5) and the bulk (300 ± 30) were found to be almost the same within error. The bulk modulus for the γ phase was found to decrease with decreasing particle size between 10 and 3.2 nm particle size. Values for the ambient pressure molar volume were found within 1% to be: 33.0 cm3 mol⁻¹ for bulk γ-Fe₂O₃; 32.8 cm³ mol⁻¹ for 7 nm diameter γ-Fe₂O₃ nanocrystals; 30.7 cm³ mol⁻¹ for bulk α-Fe₂O₃; and 30.6 cm³ mol⁻¹ for α-Fe₂O₃ nanocrystals.6 page(s

Size dependence of the pressure-induced gamma to alpha structuraltransition in iron oxide nanocrystals

The size trend for the pressure-induced gamma-Fe2O3(maghemite) to alpha-Fe2O3 (hematite) structural phase transition in nanocrystals has been observed. The transition pressure was found to increase with decreasing nanocrystal size: 7 nm nanocrystals transformed at 272GPa, 5 nm at 343GPa and 3 nm at 372GPa. Annealing of a bulk sample of gamma-Fe2O3 was found to reduce the transition pressure from 352 to242GPa. The bulk modulus was determined to be 2626GPa for 7 nm nanocrystals of gamma-Fe2O3, which is significantly higher than for the value of 1906 GPa that we measured for bulk samples. For alpha-Fe2O3, the bulk moduli for 7 nm nanocrystals (3365) and bulk (30030) were found to be almost the same within error. The bulk modulus for the gamma phase was found to decrease with decreasing particle size between 10 and 3.2 nm particle size. Values for the ambient pressure molar volume were found within 1 percent to be: 33.0 cm3/mol for bulk gamma-Fe2O3, 32.8 cm3/mol for 7 nm diameter gamma-Fe2O3 nanocrystals, 30.7 cm3/mol for bulk alpha-Fe2O3 and 30.6 cm3/mol for alpha-Fe2O3 nanocrystals

Epitope discovery in West Nile virus infection: Identification and immune recognition of viral epitopes

Cytotoxic T lymphocytes (CTL) play an important role in the control and elimination of infection by West Nile virus (WNV), yet the class I human leukocyte antigen (HLA)-presented peptide epitopes that enable CTL recognition of WNV-infected cells remain uncharacterized. The goals of this work were first to discover the peptide epitopes that distinguish the class I HLA of WNV-infected cells and then to test the T cell reactivity of newly discovered WNV epitopes. To discover WNV-immune epitopes, class I HLA was harvested from WNV (NY99 strain)-infected and uninfected HeLa cells. Then peptide epitopes were eluted from affinity-purified HLA, and peptide epitopes from infected and uninfected cells were comparatively mapped by mass spectroscopy. Six virus-derived peptides from five different viral proteins (E, NS2b, NS3, NS4b, and NS5) were discovered as unique to HLA-A*0201 of infected cells, demonstrating that the peptides sampled by class I HLA are distributed widely throughout the WNV proteome. When tested with CTL from infected individuals, one dominant WNV target was apparent, two epitopes were subdominant, and three demonstrated little CTL reactivity. Finally, a sequence comparison of these epitopes with the hundreds of viral isolates shows that HLA-A*0201 presents epitopes derived from conserved regions of the virus. Detection and recovery from WNV infection are therefore functions of the ability of class I HLA molecules to reveal conserved WNV epitopes to an intact cellular immune system that subsequently recognizes infected cells