The interleukin-2 receptor α chain (IL-2Rα) is potently induced by antigens, mitogens, and certain cytokines that include IL-2 itself. This induction leads to the formation of high affinity IL-2 ...receptors when IL-2Rα is co-expressed with the β (IL-2Rβ) and γ (γc) chains of this receptor. We investigated the signaling pathways mediating IL-2-induced IL-2Rα mRNA expression using 32D myeloid progenitor cells stably transfected with either wild type IL-2Rβ or mutants of IL-2Rβ containing tyrosine to phenylalanine substitutions. Of the six cytoplasmic tyrosines in IL-2Rβ, we have found that only the two tyrosines that mediate Stat5 activation (Tyr-392 and Tyr-510) contribute to IL-2-induced IL-2Rα gene expression and that either tyrosine alone is sufficient for this process. Interestingly, the IL-7 receptor contains a tyrosine (Tyr-429)-based sequence resembling the motifs encompassing Tyr-392 and Tyr-510 of IL-2Rβ. Further paralleling the IL-2 system, IL-7 could activate Stat5 and drive expression of IL-2Rα mRNA in 32D cells transfected with the human IL-7R. However, IL-3 could not induce IL-2Rα mRNA in 32D cells, despite its ability to activate Stat5 via the endogenous IL-3 receptor. Moreover, the combination of IL-3 and IL-2 could not “rescue” IL-2Rα mRNA expression in cells containing an IL-2Rβ mutant with phenylalanine substitutions at Tyr-392 and Tyr-510. These data suggest that Tyr-392 and Tyr-510 couple to an additional signaling pathway beyond STAT protein activation in IL-2-mediated induction of the IL-2Rα gene.
Proteomics technologies are often used for the identification of protein targets of the immune system. Here, we discuss the immunoproteomics technologies used for the discovery of autoantigens in ...autoimmune diseases where immune system dysregulation plays a central role in disease onset and progression. These autoantigens and associated autoantibodies can be used as potential biomarkers for disease diagnostics, prognostics and predicting/monitoring drug responsiveness (theranostics). Here, we compare a variety of methods such as mass spectrometry (MS)-based serological proteome analysis (SERPA), antibody mediated identification of antigens (AMIDA), circulating immune complexome (CIC) analysis, surface enhanced laser desorption/ionization-time of flight (SELDI-TOF), nucleic acid based serological analysis of antigens by recombinant cDNA expression cloning (SEREX), phage immunoprecipitation sequencing (PhIP-seq) and array-based immunoscreening (proteomic microarrays), luciferase immunoprecipitation systems (LIPS), nucleic acid programmable protein array (NAPPA) methods. We also review the relevance of immunoproteomic data generated in the last 10 years, with a focus on the aforementioned MS based methods.
Abstract
Objectives
The 2016 ACR-EULAR Response Criteria for JDM was developed as a composite measure with differential weights of six core set measures (CSMs) to calculate a Total Improvement Score ...(TIS). We assessed the contribution of each CSM, representation of muscle-related and patient-reported CSMs towards improvement, and frequency of CSM worsening across myositis response criteria (MRC) categories in validation of MRC.
Methods
Data from JDM patients in the Rituximab in Myositis trial (n = 48), PRINTO JDM trial (n = 139), and consensus patient profiles (n = 273) were included. Observed vs expected CSM contributions were compared using Sign test. Characteristics of MRC categories were compared by Wilcoxon tests with Bonferroni adjustment. Spearman correlation of changes in TIS and individual CSMs were examined. Agreement between physician-assessed change and MRC categories was evaluated by weighted Cohen’s kappa.
Results
Of 457 JDM patients with IMACS CSMs and 380 with PRINTO CSMs, 9–13% had minimal, 19–23% had moderate and 41–50% had major improvement. The number of improved and absolute percentage change of CSMs increased by MRC improvement level. Patients with minimal improvement by MRC had a median of 0–1 CSM worsened, and those with moderate/major improvement had a median of zero worsening CSMs. Of patients improved by MRC, 94–95% had improvement in muscle strength and 93–95% had improvement in ≥1 patient-reported CSM. IMACS and PRINTO CSMs performed similarly. Physician-rated change and MRC improvement categories had moderate-to-substantial agreement (Kappa 0.5–0.7).
Conclusion
The ACR-EULAR MRC perform consistently across multiple studies, supporting its further use as an efficacy end point in JDM trials.
CIS is a c ytokine- i nduced S H2-containing protein that was originally cloned as an interleukin (IL)-3-inducible gene. CIS is known to associate with the
IL-3 receptor β chain and erythropoietin ...receptor and to inhibit signaling mediated by IL-3 and erythropoietin. We now demonstrate
that CIS also interacts with the IL-2 receptor β chain (IL-2Rβ). This interaction requires the A region of IL-2Rβ (residues
313â382), which also mediates the association of IL-2Rβ with Lck and Jak3. Correspondingly, CIS inhibits functions associated
with both of these kinases: Lck-mediated phosphorylation of IL-2Rβ and IL-2-mediated activation of Stat5. Thus, we demonstrate
that CIS can negatively control at least two independent IL-2 signaling pathways. Although a functional SH2 binding domain
of CIS was not required for its interaction with IL-2Rβ in vitro , its phosphotyrosine binding capability was essential for the inhibitory action of CIS. On this basis, we have generated
a mutant form of CIS protein with an altered SH2 domain that acts as a dominant negative and should prove useful in further
understanding CIS action.
The interleukin-2 receptor α chain (IL-2Rα) is potently induced by antigens, mitogens, and certain cytokines that include
IL-2 itself. This induction leads to the formation of high affinity IL-2 ...receptors when IL-2Rα is co-expressed with the β
(IL-2Rβ) and γ (γ c ) chains of this receptor. We investigated the signaling pathways mediating IL-2-induced IL-2Rα mRNA expression using 32D
myeloid progenitor cells stably transfected with either wild type IL-2Rβ or mutants of IL-2Rβ containing tyrosine to phenylalanine
substitutions. Of the six cytoplasmic tyrosines in IL-2Rβ, we have found that only the two tyrosines that mediate Stat5 activation
(Tyr-392 and Tyr-510) contribute to IL-2-induced IL-2Rα gene expression and that either tyrosine alone is sufficient for this
process. Interestingly, the IL-7 receptor contains a tyrosine (Tyr-429)-based sequence resembling the motifs encompassing
Tyr-392 and Tyr-510 of IL-2Rβ. Further paralleling the IL-2 system, IL-7 could activate Stat5 and drive expression of IL-2Rα
mRNA in 32D cells transfected with the human IL-7R. However, IL-3 could not induce IL-2Rα mRNA in 32D cells, despite its ability
to activate Stat5 via the endogenous IL-3 receptor. Moreover, the combination of IL-3 and IL-2 could not ârescueâ IL-2Rα mRNA
expression in cells containing an IL-2Rβ mutant with phenylalanine substitutions at Tyr-392 and Tyr-510. These data suggest
that Tyr-392 and Tyr-510 couple to an additional signaling pathway beyond STAT protein activation in IL-2-mediated induction
of the IL-2Rα gene.
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