RESUMEN
In the two decades since the invention of laser-based super resolution microscopy this family of technologies has revolutionised the way life is viewed and understood. Its unparalleled resolution, speed, and accessibility makes super resolution imaging particularly useful in examining the highly complex and dynamic immune system. Here we introduce the super resolution technologies and studies that have already fundamentally changed our understanding of a number of central immunological processes and highlight other immunological puzzles only addressable in super resolution.
Asunto(s)
Técnicas Inmunológicas/instrumentación , Microscopía Confocal/métodos , Imagen Individual de Molécula/métodos , Animales , Linaje de la Célula , Diseño de Equipo , Recuperación de Fluorescencia tras Fotoblanqueo , Humanos , Sistema Inmunológico/citología , Microscopía Confocal/instrumentación , Microscopía Fluorescente/instrumentación , Microscopía Fluorescente/métodos , Receptores de Antígenos/ultraestructura , Receptores Inmunológicos/ultraestructura , Imagen Individual de Molécula/instrumentaciónRESUMEN
Over the last decade, live cell imaging has revealed the surprisingly complex orchestration of antigen receptor signalling at the immunological synapse. The imaging studies showed that one of the earliest steps in antigen receptor activation is the formation of submicroscopic clusters, which regulate the early signalling events. However, the molecular mechanisms operating inside these microclusters have remained beyond the resolution of optical microscopy. Recent development of imaging techniques that approach molecular resolution in intact cells offers a first view of the molecular processes inside these structures. Here I review the contributions of molecular imaging of the immunological synapse to our understanding of antigen receptor clustering, binding to antigens, and recruitment of signalling molecules. Finally, I provide an outlook on the future prospects of this rapidly advancing technology.
Asunto(s)
Imagen Molecular , Receptores de Antígenos/fisiología , Transducción de Señal , Transferencia Resonante de Energía de Fluorescencia , Sinapsis Inmunológicas/fisiología , Sinapsis Inmunológicas/ultraestructura , Receptores de Antígenos/ultraestructuraRESUMEN
We recently have identified an antigen receptor in sharks called NAR (new or nurse shark antigen receptor) that is secreted by splenocytes but does not associate with Ig light (L) chains. The NAR variable (V) region undergoes high levels of somatic mutation and is equally divergent from both Ig and T cell receptors (TCR). Here we show by electron microscopy that NAR V regions, unlike those of conventional Ig and TCR, do not form dimers but rather are independent, flexible domains. This unusual feature is analogous to bona fide camelid IgG in which modifications of Ig heavy chain V (VH) sequences prevent dimer formation with L chains. NAR also displays a uniquely flexible constant (C) region. Sequence analysis and modeling show that there are only two types of expressed NAR genes, each having different combinations of noncanonical cysteine (Cys) residues in the V domains that likely form disulfide bonds to stabilize the single antigen-recognition unit. In one NAR class, rearrangement events result in mature genes encoding an even number of Cys (two or four) in complementarity-determining region 3 (CDR3), which is analogous to Cys codon expression in an unusual human diversity (D) segment family. The NAR CDR3 Cys generally are encoded by preferred reading frames of rearranging D segments, providing a clear design for use of preferred reading frame in antigen receptor D regions. These unusual characteristics shared by NAR and unconventional mammalian Ig are most likely the result of convergent evolution at the molecular level.