An induced fit hypothesis for antigen recognition by T lymphocytes: a role for specific antigen retention structures on antigen-presenting cells

The nature of T lymphocyte recognition of foreign antigens is not known, despite recent advances in elucidating the cellular structures that may be involved in the specific interactions. The central difficulty in this process is that T cells respond to foreign antigen only in the context of major histocompatibility complex (MHC) antigens expressed by another antigen-presenting cell. In addition, T cells that interact with class II MHC antigens do not bind foreign protein antigens in their native form, but seem to recognize only proteolytic peptide fragments as the relevant antigen. The simplest explanation for these observations is that the class II MHC antigens themselves bind antigenic peptides to form the appropriate determinant that interacts with the antigen-specific T cell receptor. However, to date no such antigenic complex has been found with MHC antigens despite rigorous attempts at their demonstration. One alternative explanation described here is that there is no preexisting foreign antigen-MHC antigen complex prior to interaction with T cells, and it is the T cells that cause the two moieties to become associated for recognition by a single antigen-specific T cell receptor. Central to this mechanism is that foreign antigenic peptides must be associated with specific antigen retention structures (SARS) expressed by antigen-presenting cells which retain and protect the peptide on the cell surface. These SARS, upon interaction with T cell membrane moieties, would subsequently associate with MHC antigens. A hypothesis to describe this mechanism is developed to account for published observations of antigen processing by antigen-presenting cells and T cell antigen recognition, and makes several predictions that are experimentally testable. This mechanism is also generally applicable to other cellular interactions in which soluble peptide mediators may become associated with surface components of one cell type, and this newly formed complex is in turn recognized by a receptor on a second cell type to deliver functional signals.     

Heterogeneity of the cellular sources of restorative processes in vertebrates

Possible cell sources for restorative processe in adult vertebrates have been considered: (1) differentiated cells undergoing a certain reconstruction after the organ damage; (2) reserve of little-differentiated cells which can be represented by heterogenous populations. Experimental data obtained on a few most studied models of restoration of the damaged organs or their parts have been analyzed. Each particular restorative process can be realized by cell populations which differ from each other by a number of features: origin, potencies, way of formation, localization of cambium, etc. Participation of heterogenous cell populations is most pronounced upon stimulation of restorative processes and in the extreme conditions. Specific involvement of every possible cell source in a particular case of restoration depends on the organism's properties, peculiarities of the damaged organ, as well as on conditions of damage. The existence of diverse cell sources provides for certain reserves of restoration what appears to be indispensable for reliability of restorative processes in varying conditions of damage.     

Heterogeneity and inaccuracy in protein structures solved by X-ray crystallography

Proteins are dynamic molecules, exhibiting structural heterogeneity in the form of anisotropic motion and discrete conformational substates, often of functional importance. In protein structure determination by X-ray crystallography, the observed diffraction pattern results from the scattering of X-rays by an ensemble of heterogeneous molecules, ordered and oriented by packing in a crystal lattice. The majority of proteins diffract to resolutions where heterogeneity is difficult to identify and model, and are therefore approximated by a single, average conformation with isotropic variance. Here we show that disregarding structural heterogeneity introduces degeneracy into the structure determination process, as many single, isotropic models exist that explain the diffraction data equally well. The large differences among these models imply that the accuracy of crystallographic structures has been widely overestimated. Further, it suggests that analyses that depend on small differences in the relative positions of atoms may be flawed.     

Optical imaging of nanoscale cellular structures

Visualization of subcellular structures and their temporal evolution is of utmost importance to understand a vast range of biological processes. Optical microscopy is the method of choice for imaging live cells and tissues; it is minimally invasive, so processes can be observed over extended periods of time without generating artifacts due to intense light irradiation. The use of fluorescence microscopy is advantageous because biomolecules or supramolecular structures of interest can be labeled specifically with fluorophores, so the images reveal information on processes involving only the labeled molecules. The key restriction of optical microscopy is its moderate resolution, which is limited to about half the wavelength of light (～200 nm) due to fundamental physical laws governing wave optics. Consequently, molecular processes taking place at spatial scales between 1 and 100 nm cannot be studied by regular optical microscopy. In recent years, however, a variety of super-resolution fluorescence microscopy techniques have been developed that circumvent the resolution limitation. Here, we present a brief overview of these techniques and their application to cellular biophysics.     

Heterogeneity in the cellular protein profiles ofClostridium botulinum types A and B

Summary Profiles of cellular proteins from 11 type A and 9 type B strains ofClostridium botulinum were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Cellular protein profiles exhibited considerable heterogeneity among strains of the same biotype, as well as among strains of different biotypes. This study also demonstrated the reliability and usefulness of this rapid and inexpensive procedure to determine the variation which may occur amongC. botulinum and related species.References to brand or firm names do not constitute endorsement by the U.S. Department of Agriculture over others of a similar nature not mentioned.     

Particle retention in suspension-feeding fish after removal of filtration structures

The suspension-feeding cichlids Oreochromis aureus (blue tilapia) and Oreochromis esculentus (ngege tilapia) are able to selectively retain small food particles. The gill rakers and microbranchiospines of these species have been assumed to function as filters. However, surgical removal of these oral structures, which also removed associated mucus, did not significantly affect the total number of 11–200 μm particles ingested by the fish. This result supports the hypothesis that the branchial arch surfaces themselves play an important role in crossflow filtration. Both species selectively retained microspheres greater than 50 μm with gill rakers and microbranchiospines intact as well as removed, demonstrating that neither these structures nor mucus are necessary for size selectivity to occur during biological crossflow filtration. After removal of the gill rakers and microbranchiospines, O. esculentus retained significantly more microspheres 51–70 μm in diameter and fewer 91–130 μm microspheres compared to retention with intact structures, but the particle size selectivity of O. aureus was not affected significantly. These results support conclusions from previous computational fluid dynamics simulations indicating that particle size can have marked effects on particle trajectory and retention inside the fish oropharyngeal cavity during crossflow filtration. The substantial inter-individual variability in particle retention by suspension-feeding fish is an unexplored area of research with the potential to increase our understanding of the factors influencing particle retention during biological filtration.     

Proteomics of organelles and large cellular structures

The mass-spectrometry-based identification of proteins has created opportunities for the study of organelles, transport intermediates and large subcellular structures. Traditional cell-biology techniques are used to enrich these structures for proteomics analyses, and such analyses provide insights into the biology and functions of these structures. Here, we review the state-of-the-art proteomics techniques for the analysis of subcellular structures and discuss the biological insights that have been derived from such studies.     

Observations on cellular structures of Porphyridium cruentum

The cellular structure of Porphyridium cruentum was studied with both light and electron microscope. The photosynthetic plastid in this red alga was found to be structurally similar to that in the Chlorophyceae and higher green plants. The phycobilins, as well as the chlorophyll, seem to be associated with the lamellae of the plastid. The pyrenoid, a region of low lamellar density, contains no tubules, and does not appear to function in synthesis or storage of reserve material. Grains of floridean starch are located in the cytoplasm, outside the plastid. Typical mitochondrial organelles were not observed. The nucleus is eccentric, and contains a nucleolus located on the inner face of the nucleus, nearest the plastid. The schedule for staining the nucleus is given in detail. Other cell structures (sheath, dictyosomes, etc.) are described. Growing cells in light of intensity leads to disruption of the parallel arrangement of the lamellar characteristic of cells grown in moderate light.     

Heterogeneity of mouse Thy 1.2 antigen expression revealed by monoclonal antibodies

A different sensitivity of T cells from C57B1/6 and DBA/2 mice to treatment with the monoclonal anti-Thy 1.2 F7D5 serum as compared with a conventional alloantiserum is reported. Depletion of T helper cells, Con A-, PHA-, MLC-, and GVH-reactive cells from a DBA/2 or C57B1/6 spleen cell population was readily achieved with the conventional alloserum. In contrast, the F7D5 antiserum abolished all T functions studied in C57B1/6 spleen cells whereas it was totally or partially ineffective on DBA/2 spleen cells when T helper, MLC, or GVH reactivity were assayed. It did however eliminate the capacity of DBA/2 spleen cells to respond to stimulation with Con A or PHA. Analysis in an Ortho-Cytofluorograf of thymocytes and sIg^? lymphocytes labeled with either GAMB-F or F7D5 + RAM Ig-F showed no difference at the level of the thymocytes: Thy 1.2 antigen as revealed by either GAMB or F7D5 is similarly expressed in the two mouse strains. The fluorescence profiles of splenic T lymphocytes indicated a reduced representation per unit cell basis of the Thy 1.2 antigenic determinant recognized by F7D5 in DBA/2 mice. Moreover, this same determinant is expressed in only 70% of all Thy 1.2-positive cells detected in DBA/2 sIg^? population. This implies that, in DBA/2 mice, maturation of T cells is accompanied by a complete or partial loss of the F7D5 Thy 1.2 determinant and that T helper functions and MLC and GVH reactivity are mediated by T cells which express little or none of this F7D5 Thy 1.2 determinant.     

Tissue localization and retention of antigen in relation to the immune response

Antigen persists for months or even years in lymphoid tissues of immune animals and this antigen is believed to participate in the induction and maintenance of B-cell memory as well as in the maintenance of serum antibody levels. In the present report we describe the phenomenon of antigen localization and long-term retention on mouse follicular dendritic cells (FDCs). The antigens used were injected in the hind footpads of immune mice and the popliteal lymph nodes were the lymphoid organs generally studied. In addition to presenting the morphological features of mouse FDCs, we report the results of a study of the mechanism of antigen migration from the site of initial localization in the lymph node subcapsular sinus to the regions of follicular retention in the cortex. The migration was followed by light and electron microscopy. The results support the concepts that immune complexes are trapped in the subcapsular sinus and are transported by a group of nonphagocytic cells to follicular regions. The mechanism of transport may involve either migration of pre-FDCs with a concomitant maturation into FDCs, or cell-to-cell transport utilizing dendritic cell processes and membrane fluidity; or a combination of the two mechanisms may be in operation.     

4Pi microscopy of quantum dot-labeled cellular structures

The most prominent restrictions of fluorescence microscopy are the limited resolution and the finite signal. Established conventional, confocal, and multiphoton microscopes resolve at best approximately 200nm in the focal plane and only 500nm in depth. Additionally, organic fluorophores and fluorescent proteins are bleached after 10(4)-10(5) excitation cycles. To overcome these restrictions, we synergistically combine the 3- to 7-fold improved axial resolution of 4Pi microscopy with the greatly enhanced photostability of semiconductor quantum dots. Co-localization studies of immunolabeled microtubules and mitochondria demonstrate the feasibility of this approach for routine biological measurements. In particular, we visualize the three-dimensional entanglement of the two networks with unprecedented detail.     

From sensing cellular sterols to assembling sensory structures

The genome of the nematode C. elegans is peppered with novel genes belonging to a superfamily whose members function in cellular cholesterol homeostasis (Niemann-Pick C1) and Hedgehog signal transduction (Patched) and biogenesis (Dispatched). In this issue of Developmental Cell and an analysis of a pair of Patched- and Dispatched-related proteins in C. elegans extends the superfamily's repertoire to include the formation of tubular organs.     

Quantitative motion analysis and visualization of cellular structures

The availability of cellular markers tagged with the green fluorescent protein (GFP) has recently allowed a large number of cell biological studies to be carried out in live cells, thereby addressing the dynamic organization of cellular structures. Typically, microscopes capable of video recording are used to generate time-resolved data sets. Dynamic imaging data are complex and often difficult to interpret by pure visual inspection. Therefore, specialized image processing methods for object detection, motion estimation, visualization, and quantitation are required. In this review, we discuss concepts for automated analysis of multidimensional image data from live cell microscopy and their application to the dynamics of cell nuclear subcompartments.     

Heterogeneity in antigen processing by different types of antigen-presenting cells. Effect of cell culture on antigen processing ability.

The ability of normal B cells, peritoneal macrophages, and splenic APC to process and present OVA to a panel of T-T hybridomas with different specificities was investigated. In all cases, B cells were less efficient than unfractionated splenocytes in presenting OVA or its peptides. However, when the presentation of native Ag was compared to the presentation of peptides, it was obvious that there were marked differences in the ability of these two APC populations to generate different epitopes from OVA. Leupeptin inhibits the processing of selected epitopes from native OVA differently when it was presented by spleen cells or B cells, suggesting that these two APC populations differ in their protease content. The effect of in vitro culture on the ability of splenic and peritoneal APC to process OVA was also investigated. Native OVA presentation by macrophages and spleen cells was affected by in vitro culture, more for some epitopes than for other epitopes. In contrast, presentation of exogenous peptides by paraformaldehyde-fixed APC was either not affected by previous culturing for 3 days, or very much improved. Altogether, these data demonstrate that different epitopes on the same protein may be independently and differentially processed by B cells and spleen cells. Furthermore, the precise peptides that are produced may vary with the physiologic state of the APC.     

Transfer factor and cellular reactivity to varicella-zoster antigen in childhood leukemia

The present studies were designed to evaluate possible efficacy of specific human transfer factor (TF) in preventing or attenuating varicella-zoster (VZ) infection in children with acute lymphocytic leukemia (ALL). TF was prepared following leukapheresis of adult donors who were convalescing from chicken pox. Recipients were VZ seronegative children with ALL, 12 in remission and 3 in relapse; a single injection of TF was given equivalent to 10⁸ lymphocytes per 7 kg body wt. The following VZ-specific parameters were measured: lymphocyte blastogenesis, cytotoxicity and leukocyte inhibitory factor (LIF) production, and indirect fluorescent and CF antibody titers. No patients in relapse converted immune response while $\text{10}\text{12}$ in remission developed positive reactivity in at least one assay of cell-mediated immunity (CMI); $\text{3}\text{12}$ were positive in all three parameters of CMI and $\text{8}\text{12}$ in two assays. Cytotoxicity was the most consistently positive test following TF administration. No patients developed VZ antibody. Seventeen months follow-up has demonstrated persistent cellular immunity.     

Heterogeneity of cellular expression of arachidonate 15-lipoxygenase: implications for biological activity

Comparison of relative arachidonate 15-lipoxygenase activities in different types of human cell reveals striking differences in levels of product generation. Among available human cell types, the tracheal epithelial cell and the eosinophil have markedly higher levels of activity than other cell types with reported activity. Theories of functional significance for this enzymatic pathway will need to account for the selective expression of high levels of activity in a highly limited number of cell types.