Immune proteasomes and immunity

A lot of facts that require understanding have been accumulated since immune proteasomes were discovered and their relationship with the immune response was established. For example, why are immune proteasomes present in all studied mammalian organs and tissues, including nonlymphoid tissues? What is responsible for differences in the ratio of immune to constitutive proteasomes in different organs? Are the functions of immune proteasomes related to the T-cell immune response alone, as was shown initially, or not? Are immune proteasomes formed simultaneously in different organs during ontogenesis? An attempt is made in this review to answer these and other related questions.     

Formation of immune proteasomes and development of immune system in ontogenesis of mammals

Current concepts of the structure of immune proteasomes and their role in immune response have been considered. The main attention has been paid to the formation of immune proteasomes in secondary lymphoid and nonlymphoid organs during ontogenesis of mammals. The causes of ineffective formation of immune system in early postnatal development have been discussed.     

Formation of immune proteasomes and development of the immune system in mammalian ontogenesis

Current concepts of the structure of immune proteasomes and their role in immune response have been considered. The main attention has been paid to the formation of immune proteasomes in secondary lymphoid and nonlymphoid organs during ontogenesis of mammals. The causes of ineffective formation of immune system in early postnatal development have been discussed.     

Exclusion of immune proteasomes from mouse ascitic carcinoma Krebs-II cells

Pools of 26S and 20S proteasomes were studied in the spleen, liver, lung, and ascitic carcinoma Krebs-II of mouse. Western blotting demonstrated that the pool of 26S proteasomes in ascitic carcinoma Krebs-II was twice that in control lung cells and did not significantly differ by total 26S proteasome quantities from the spleen and liver. At the same time, the level of immune subunit LMP7 was 12 times lower in it compared to lung proteasomes and 4–5 times lower compared to spleen and liver proteasomes. Immune subunit LMP2 was undetectable by this technique in the ascitic carcinoma in contrast to the lung, spleen, and liver. All immune subunits in the studied organs and ascitic carcinoma Krebs-II are components of 26S but not 20S proteasomes.     

Effects of Dietary Selenium on Selenoprotein W Gene Expression in the Chicken Immune Organs

Selenoprotein W (SelW) is expressed in the immune systems of mammals. However, its pattern of expression in the immune organs of birds is still unclear. To investigate the distribution of SelW and effects of dietary Se levels on the SelW mRNA expression in the immune organs of birds, 1-day-old male chickens were fed either a commercial diet or an Se-supplemented diet containing 0.601, 1.058, 1.514, or 2.427?mg Se per kilogram, and 1.0, 2.0, 3.0 or 5.0?mg sodium selenite per kilogram for 90?days. The immune organs (spleen, thymus, and bursa of Fabricius) were collected and examined for Se content and SelW mRNA levels. The mRNA expression of SelW was detected in all the tissues. Although Se content was the highest in the spleen, the remarkable stability of the SelW mRNA level was observed in this organ during different times of dietary Se supplementation. Se-supplemented diet can make the SelW expression levels higher within a certain range in thymus and bursa of Fabricius. The present study demonstrates that SelW is widely expressed in immune organs of birds and that Se-supplementation of the feed increases SelW expression in the thymus and the bursa of Fabricius.     

Peculiarities of proteasome pool formation in rat spleen and liver during postnatal development

Changes in the specific activity and amounts of 26S and 20S proteasome pools in rat spleen and liver during postnatal development and appearance in them of immune subunits were studied. Two decreases in chymotrypsin-like activity of the proteasome pools were recorded during the first three weeks after birth. The activity minimum fell on the 11th and 19th days, and the first decrease was more prolonged and pronounced than the second. The decrease in the specific activity of the 26S proteasome pools was associated with a reduction of their quantity. The 20S proteasome pools displayed no such decreases. Noticeable quantities of immune subunits LMP7 and LMP2 were revealed by Western blotting in the spleen on the 7th day and on the 19th day in the liver, concurrently with the beginning of the decrease in the proteasome activity. It was concluded that during the first three weeks of postnatal development the proteasome pools in rat spleen and liver were replaced twice, and in the spleen (a lymphoid organ) a qualitatively new pool containing immune subunits appeared nearly two weeks earlier than in the liver (a non-lymphoid organ). The appearance of immune proteasomes in different organs and tissues during some weeks after birth seems to explain the immune system inefficiency during embryogenesis and early postnatal development.     

Contrasting proteome biology and functional heterogeneity of the 20 S proteasome complexes in mammalian tissues

The 20 S proteasome complexes are major contributors to the intracellular protein degradation machinery in mammalian cells. Systematic administration of proteasome inhibitors to combat disease (e.g. cancer) has resulted in positive outcomes as well as adversary effects. The latter was attributed to, at least in part, a lack of understanding in the organ-specific responses to inhibitors and the potential diversity of proteomes of these complexes in different tissues. Accordingly, we conducted a proteomic study to characterize the 20 S proteasome complexes and their postulated organ-specific responses in the heart and liver. The cardiac and hepatic 20 S proteasomes were isolated from the same mouse strain with identical genetic background. We examined the molecular composition, complex assembly, post-translational modifications and associating partners of these proteasome complexes. Our results revealed an organ-specific molecular organization of the 20 S proteasomes with distinguished patterns of post-translational modifications as well as unique complex assembly characteristics. Furthermore, the proteome diversities are concomitant with a functional heterogeneity of the proteolytic patterns exhibited by these two organs. In particular, the heart and liver displayed distinct activity profiles to two proteasome inhibitors, epoxomicin and Z-Pro-Nle-Asp-H. Finally, the heart and liver demonstrated contrasting regulatory mechanisms from the associating partners of these proteasomes. The functional heterogeneity of the mammalian 20 S proteasome complexes underscores the concept of divergent proteomes among organs in the context of an identical genome.     

Naturally occurring genetic variation in the age-specific immune response of Drosophila melanogaster

Immunosenescence, the age‐related decline in immune response, is a well‐known consequence of aging. To date, most studies of age‐related changes in immune response focused on the cellular and physiological bases of this decline; we have virtually no understanding of the genetic basis of age‐related changes in the immune system or if indeed such control exists. We used 25 chromosome substitution lines of Drosophila melanogaster derived from a natural population to address three questions: (i) How is the function of the innate immune system influenced by age? (ii) Is there a genetic basis for phenotypic variation in immune response at different ages? (iii) Is there a genetic basis for differences in the way that age influences the immune function? Virgin females from each line were assayed for immune response using clearance of infection with Escherichia coli at 1 and 4 weeks of age. We found significant genetic variation among lines in immune response at each age. Unexpectedly, when averaged across all lines, the immune response actually improved with age. However, there was significant variation in the effect of age on immune response with 11 lines showing improvement, nine lines showing no change and five exhibiting a decline with age. There was no genetic correlation of immune response across ages suggesting that different loci contribute to variation in immune response at each age. The genetic component of the variation in immune response increased with age, a pattern predicted by the mutation accumulation model of senescence. However, this increase in variation resulted in part from the improvement of the immune response in some lines with age. Thus the observed changes in genetic variation in immune function with age are not entirely explained by the mutation accumulation model.     

Different proteasome subtypes in a single tissue exhibit different enzymatic properties

It is concluded from many experiments that mammalian tissues and cells must contain a heterogeneous population of 20 S proteasome complexes. We describe the purification and separation by chromatographic procedures of constitutive 20 S proteasomes, 20 S immuno-proteasomes and intermediate-type 20 S proteasomes from a given tissue. Our data demonstrate that each of these three groups comprises more than one subtype and that the relative ratios of the subtypes differ between different rat tissues. Thus, six subtypes could be identified in rat muscle tissue. Subtypes I and II are constitutive proteasomes, while subtypes V and VI comprise immuno-proteasomes. Subtypes III and IV belong to a group of intermediate-type proteasomes. The subtypes differ with regard to their enzymatic characteristics. Subtypes I-III exhibit high chymotrypsin-like activity and high peptidylglutamylpeptide hydrolysing activity, while these activities are depressed in subtypes IV-VI. In contrast, trypsin-like activity of subtypes IV-VI is enhanced in comparison to subtypes I-III. Importantly, the subtypes also differ in their preferential cleavage site usage when tested by digestion of a synthetic 25mer polypeptide substrate. Therefore, the characteristics of proteasomes purified from tissues or cells represent the average of the different subtype activities which in turn may have different functions in vivo.     

CCR7 coordinates the primary immune response by establishing functional microenvironments in secondary lymphoid organs.

The proper function of immune surveillance requires well-coordinated mechanisms in order to guide the patrolling immune cells through peripheral tissues and into secondary lymphoid organs. Analyzing gene-targeted mice, we identified the chemokine receptor CCR7 as an important organizer of the primary immune response. CCR7-deficient mice show severely delayed kinetics regarding the antibody response and lack contact sensitivity and delayed type hypersensitivity reactions. Due to the impaired migration of lymphocytes, these animals reveal profound morphological alterations in all secondary lymphoid organs. Upon activation, mature skin dendritic cells fail to migrate into the draining lymph nodes. Thus, in order to bring together lymphocytes and dendritic cells to form the characteristic microarchitecture of secondary lymphoid organs, CCR7 is required to rapidly initiate an adoptive immune response.     

Proteasomes on thyroid tissue allotransplantation under induction of donor-specific tolerance in rats

Proteasomes in the liver of August rats (RT1^c) were investigated 30 days after allotransplantation of Wistar rat (RT1^u) thyroid tissue under renal capsule with/without induction of donor-specific tolerance by donor splenocyte intraportal administration. The levels of total proteasome pool, immune proteasomes containing subunits LMP2 and/or LMP7, and proteasome regulators 19S and 11S were defined. Intact and sham-operated August rats were used as control groups. The level of all immune proteasome forms and 11S regulator increased while the level of the total proteasome pool and 19S regulator decreased in the liver of experimental animals compared to the control groups, which indicated changes of liver functional state after transplantation. The 19S/11S ratio increased in the liver of nontolerant rats compared to tolerant animals. In the liver of tolerant rats with accepted grafts, the number of mononuclear cells expressing the immune subunit LMP2 greatly increased in comparison with control and nontolerant animals. Study of accepted grafts showed an increase in the ratio of LMP2/LMP7 immune subunits and 19S/11S regulators in them, compared to the tissue replacing the rejected grafts. Immune proteasomes were almost completely absent from the control intact thyroid tissue, while 19S/11S ratio was maximal in it. Thus, the development of the immune reaction or its suppression are accompanied by a change in the balance between different proteasome forms. Immune subunit LMP7 and 11S regulator are associated with the response against donor tissue. On the contrary, immune subunit LMP2 and 19S regulator are likely to be important for the development of immune tolerance and surviving tissue functioning. Immunofluorescence assay revealed a low content of the immune proteasomes in the follicle cells. Probably, formation of antigens for the major histocompatibility complex class I molecules was impaired by the low content of immune proteasomes, which led to immunological tolerance of hormone-producing follicle cells.     

Effect of the thymus on the activity of immunoreactive peptides

Influence of the thymic gland on the biology activity of peptide mediators--cytomedins, has been studied in the content of different organs. The peptide which was obtained from the liver, spleen and muscle of sham-operated animals enhanced immune response on the sheep red cells, lengthened indexes of thromboelastography, and activated fibrinolysis after its administration to the thymectomized rats. The peptides from organs of thymectomized animals lost this ability. These findings confirm the hypothesis on the regulating function of the thymic gland in the synthesis and activity of cytomedins in the tissues.     

A Mathematical Model of the Inflammatory Response to Pathogen Challenge

The human body’s immune response to bacterial challenge, even when successful in controlling the infection, can result in negative consequences for the host, including reduced functionality of associated tissues. We present and analyze a low-dimensional mathematical model of this immune response to pathogen invasion, incorporating the coordinated actions of active immune cells, and both pro- and anti-inflammatory cytokines. The model simulates both the positive (pathogen reduction) and negative (local tissue dysfunction) effects of the immune response and includes the important role of immunologic memory in the process of a return to stasis. This differential equation-based model is sufficiently general to be applicable to a wide range of human tissues and organs.     

Immune proteasomes in the developing rat spleen

Changes in the structure of the rat spleen and the distribution of immune proteasomes in it during early postnatal development have been studied using double immunofluorescent staining of tissue sections with antibodies to the LMP7 immune proteasome subunit and to specific markers of T and B lymphocytes. It has been shown that the white pulp on postnatal day 5 is not yet colonized by lymphocytes and contains a smaller amount of immune proteasomes than the red pulp. At this stage, T and B lymphocytes concentrate mainly in the red pulp. On day 8, B lymphocytes occupy the marginal zone, while T lymphocytes aggregate into dense strands close to the white pulp. By day 18, T lymphocytes form periarteriolar sheaths in the white pulp, and the contents of immune proteasomes in the red and white pulp become equally high. An increase in the total content of immune proteasomes in the spleen on the third postnatal week was revealed in our previous study by Western blotting. In addition to T and B lymphocytes, immune proteasomes have also been revealed in other spleen cell types, probably in macrophages and reticular cells of the white pulp. Thus, the postnatal development of the spleen is associated with an increase in the contents of immune proteasomes in it.     

Immunologic 'ignorance' of vascularized organ transplants in the absence of secondary lymphoid tissue

Secondary lymphoid organs (the spleen, lymph nodes and mucosal lymphoid tissues) provide the proper environment for antigen-presenting cells to interact with and activate naive T and B lymphocytes. Although it is generally accepted that secondary lymphoid organs are essential for initiating immune responses to microbial antigens and to skin allografts, the prevailing view has been that the immune response to primarily vascularized organ transplants such as hearts and kidneys does not require the presence of secondary lymphoid tissue. The assumption has been that the immune response to such organs is initiated in the graft itself when recipient lymphocytes encounter foreign histocompatibility antigens presented by the graft's endothelial cells. In contrast to this view, we show here that cardiac allografts are accepted indefinitely in recipient mice that lack secondary lymphoid tissue, indicating that the alloimmune response to a vascularized organ transplant cannot be initiated in the graft itself. Moreover, we demonstrate that the permanent acceptance of these grafts is not due to tolerance but is because of immunologic 'ignorance'.     

Chymotrypsin-like activity and subunit composition of proteasomes in human cancers

The activity of the proteasome, a polyfunctional enzymatic complex, is known to undergo changes during cancer development. This phenomenon is probably caused by the changes in subunit composition of proteasomes. In this work, we studied the chymotrypsin-like activity of proteasomes; their subunit composition; and their association in breast cancer, head and neck squamous cell carcinoma, endometrial cancer, renal cancer, bladder cancer, stomach cancer, and colorectal cancer. The increase in proteasome activity was revealed in most cancer tissues compared with adjacent tissues, except for the renal cell carcinoma. Changes in proteasome activity in cancer tissues compared with correspondent normal tissues observed in combination with an increased expression of immune subunits and/or proteasome activator PA28β associated with activity of 20S proteasome. In breast cancer, head and neck squamous cell carcinoma, bladder cancer, stomach cancer, and colorectal cancer, we additionally found the higher expression of Rpt6 subunit of the 19S-subunit in 26S proteasome. Correlations between chymotrypsin-like proteasome activity and subunit expressions were found in human cancer tissues. Thus, we suggest that proteasome activation and changes in its subunit composition play an important role in cancer pathogenesis.     

Immunoproteasomes largely replace constitutive proteasomes during an antiviral and antibacterial immune response in the liver.

The proteasome is critically involved in the production of MHC class I-restricted T cell epitopes. Proteasome activity and epitope production are altered by IFN-gamma treatment, which leads to a gradual replacement of constitutive proteasomes by immunoproteasomes in vitro. However, a quantitative analysis of changes in the steady state subunit composition of proteasomes during an immune response against viruses or bacteria in vivo has not been reported. Here we show that the infection of mice with lymphocytic choriomeningitis virus or Listeria monocytogenes leads to an almost complete replacement of constitutive proteasomes by immunoproteasomes in the liver within 7 days. Proteasome replacements were markedly reduced in IFN-gamma(-/-) mice, but were only slightly affected in IFN-alphaR(-/-) and perforin(-/-) mice. The proteasome regulator PA28alpha/beta was up-regulated, whereas PA28gamma was reduced in the liver of lymphocytic choriomeningitis virus-infected mice. Proteasome replacements in the liver strongly altered proteasome activity and were unexpected to this extent, since an in vivo half-life of 12 days had been previously assigned to constitutive proteasomes in the liver. Our results suggest that during the peak phase of viral and bacterial elimination the antiviral cytotoxic T lymphocyte response is directed mainly to immunoproteasome-dependent T cell epitopes, which would be a novel parameter for the design of vaccines.     

Changes in proteasome pool in human papillary thyroid carcinoma development

Searching the antitumor drug targets among proteasomes, “ubiquitous” enzyme systems, may provide a new impulse to the antitumor drug discovery. In this study, changes in the proteasome pool in the development of human papillary thyroid carcinoma were determined. Proteasome activities were evaluated by hydrolysis of commercial fluorogenic peptides. Changes in the expression of the total proteasome pool, proteasome 19S activator and proteolytic constitutive subunits X(β5), Y(β1) and immune subunits LMP7 (β5i) and LMP2 (β1i) were investigated by Western blotting. The distribution of the proteasome subunits in thyroid gland cells was detected by immunohistochemistry. It was shown that the chymotrypsin- and caspase-like activities as well as the expression of the total proteasome pool, proteasome 19S activator and immune subunits increased gradually in the tumors at the T₂N₀M₀ and T₃N₀M₀ stages in comparison with the control tissues. Among the structures studied, the expression of the 19S activator and immune proteasomes, which contain the LMP2 (β1i) subunit, was enhanced to the largest degree in tumor cells. The data obtained may be implicated in a new therapeutic strategy. Taking into consideration the antitumor function of the immune proteasomes, we advance the 19S activator as the target for the development of a novel antitumor therapy.     

Adrm1, a putative cell adhesion regulating protein, is a novel proteasome-associated factor

We have identified Adrm1 as a novel component of the regulatory ATPase complex of the 26 S proteasome: Adrm1 was precipitated with an antibody to proteasomes and vice versa. Adrm1 co-migrated with proteasomes on gel-filtration chromatography and non-denaturing polyacrylamide gel electrophoresis. Adrm1 has been described as an interferon-gamma-inducible, heavily glycosylated membrane protein of 110 kDa. However, we found Adrm1 in mouse tissues only as a 42 kDa peptide, corresponding to the mass of the non-glycosylated peptide chain, and it could not be induced in HeLa cells with interferon. Adrm1 was present almost exclusively in soluble 26 S proteasomes, albeit a small fraction was membrane-associated, like proteasomes. Adrm1 was found in cells in amounts equimolar with S6a, a 26 S proteasome subunit. HeLa cells contain no pool of free Adrm1 but recombinant Adrm1 could bind to pre-existing 26 S proteasomes in cell extracts. Adrm1 may be distantly related to the yeast proteasome subunit Rpn13, mutants of which are reported to display no obvious phenotype. Accordingly, knock-down of Adrm1 in HeLa cells had no effect on the amount of proteasomes, or on degradation of bulk cell protein, or accumulation of polyubiquitinylated proteins. This indicates that Adrm1 has a specialised role in proteasome function.     

Umbilical cord fibroblasts: Could they be considered as mesenchymal stem cells?

In cell therapy protocols, many tissues were proposed as a source of mesenchymal stem cells (MSC) isolation. So far, bone marrow (BM) has been presented as the main source of MSC despite the invasive isolation procedure related to this source. During the last years, the umbilical cord (UC) matrix was cited in different studies as a reliable source from which long term ex vivo proliferating fibroblasts were isolated but with contradictory data about their immunophenotype, gene expression profile, and differentiation potential. Hence, an interesting question emerged: Are cells isolated from cord matrix (UC-MSC) different from other MSCs? In this review, we will summarize different studies that isolated and characterized UC-MSC. Considering BM-MSC as gold standard, we will discuss if UC-MSC fulfill different criteria that define MSC, and what remain to be done in this issue.