The regulation of interleukin-2-induced vimentin gene expression in a cloned T lymphocyte

Nucleotide sequencing of a 1748 base pair murine vimentin cDNA demonstrates that the 466 amino acid murine vimentin monomer possesses structural features characteristic of intermediate filament chains. Vimentin gene expression is growth- associated in the murine helper T cell clone, L2, with steady-state vimentin mRNA levels increasing 7- to 20-fold following interleukin 2 (IL2) stimulation. Accompanying a 7-fold increase in vimentin mRNA levels is a 3-fold increase in vimentin mRNA half-life, indicating that mRNA stabilization is one of several mechanisms responsible for this induction of vimentin mRNA expression. Steady-state vimentin precursor RNA levels increase 3- to 4-fold following IL2 stimulation, while vimentin precursor RNA half-life remains constant. These results indicate that precursor RNA stabilization does not contribute to the upregulation of vimentin precursor RNA expression, and suggest that IL2 induces an increase in the rate of vimentin gene transcription. The 7 to 20-fold increase in vimentin mRNA levels is paralleled by a 14-fold increase in the rate of vimentin protein synthesis. This suggests that translation is occurring, and thus that vimentin expression is regulated at least in part, by the abundance of vimentin mRNA present. In contrast to the increases in steady-state vimentin mRNA and the rate of vimentin protein synthesis, steady-state vimentin protein levels increase maximally only 1.3- to 3-fold in proliferating L2 cells. This increase in vimentin protein content is associated with recently activated and actively proliferating cells, with protein levels accumulating as the cells progress through the cell cycle. The rate of vimentin protein degradation remains relatively constant throughout L2 cell activation, active proliferation and return to quiescence, indicating that an increased rate of degradation does not account for the minimal increase in vimentin protein content. These observations may be accounted for by a transient increase in vimentin protein synthesis necessary to support L2 cell growth and/or vimentin network reorganization. Due to the transient nature of this increase, and the fact that cell division occurs every 34 h, equilibrium at this increased rate of synthesis is never reached

Mapping of the insulin-dependent diabetes locus Idd3 to a 0.35cM interval containing the Interleukin-2 gene

Currently, 16 loci that contribute to the development of IDDM in the NOD mouse have been mapped by linkage analysis. To fine map these loci, we used congenic mapping. Using this approach, we localized the Idd3 locus to a 0.35-cM interval on chromosome 3 containing the Il2 gene. Segregation analysis of the known variations within this interval indicated that only one variant, a serine-to-proline substitution at position 6 of the mature interleukin-2 (IL-2) protein, consistently segregates with IDDM in crosses between NOD and a series of nondiabetic mouse strains. These data, taken together with the immunomodulatory role of IL-2, provide circumstantial evidence in support of the hypothesis that Idd3 is an allelic variation of the Il2 gene, or a variant in strong linkage disequilibrium

Genetic control of autoimmune diabetes

By congenic strain mapping using autoimmune NOD.C57BL/6J congenic mice, we demonstrated previously that the type 1 diabetes (T1D) protection associated with the insulin-dependent diabetes (Idd)10 locus on chromosome 3, originally identified by linkage analysis, was in fact due to three closely linked Idd loci: Idd10, Idd18.1, and Idd18.3. In this study, we define two additional Idd loci--Idd18.2 and Idd18.4--within the boundaries of this cluster of disease-associated genes. Idd18.2 is 1.31 Mb and contains 18 genes, including Ptpn22, which encodes a phosphatase that negatively regulates T and B cell signaling. The human ortholog of Ptpn22, PTPN22, is associated with numerous autoimmune diseases, including T1D. We, therefore, assessed Ptpn22 as a candidate for Idd18.2; resequencing of the NOD Ptpn22 allele revealed 183 single nucleotide polymorphisms with the C57BL/6J (B6) allele--6 exonic and 177 intronic. Functional studies showed higher expression of full-length Ptpn22 RNA and protein, and decreased TCR signaling in congenic strains with B6-derived Idd18.2 susceptibility alleles. The 953-kb Idd18.4 locus contains eight genes, including the candidate Cd2. The CD2 pathway is associated with the human autoimmune disease, multiple sclerosis, and mice with NOD-derived susceptibility alleles at Idd18.4 have lower CD2 expression on B cells. Furthermore, we observed that susceptibility alleles at Idd18.2 can mask the protection provided by Idd10/Cd101 or Idd18.1/Vav3 and Idd18.3. In summary, we describe two new T1D loci, Idd18.2 and Idd18.4, candidate genes within each region, and demonstrate the complex nature of genetic interactions underlying the development of T1D in the NOD mouse model.HIF was funded by a Wellcome Trust 4-year studentship. This work was supported by Wellcome Trust Grant 091157 and JDRF International Grant 9-2011-253. Cambridge Institute for Medical Research is in receipt of a Wellcome Trust Strategic Award (100140). This work was also supported by Awards P01AI039671 (L.S.W. and J.A.T.), R01AI070544 (N.B.) and U01AI070351 (L.A.S.) from the National Institute of Allergy and Infectious Diseases at the National Institutes of Health. S.M.S. was supported by a postdoctoral fellowship from JDRF. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of Allergy and Infectious Diseases or the National Institutes of Health. The resequencing of Ptpn22 in the NOD mouse strain was performed at WTSI and was funded by the Immune Tolerance Network contract AI15416, which was sponsored by the National Institute of Allergy and Infectious Diseases, the National Institute of Diabetes and Digestive and Kidney Diseases, and the Juvenile Diabetes Research Foundation International.This is the final version of the article. It first appeared from the American Association of Immunologists via http://dx.doi.org/10.4049/jimmunol.140265

Ptpn22 and Cd2 Variations Are Associated with Altered Protein Expression and Susceptibility to Type 1 Diabetes in Nonobese Diabetic Mice

By congenic strain mapping using autoimmune NOD.C57BL/6J congenic mice, we demonstrated previously that the type 1 diabetes (T1D) protection associated with the insulin-dependent diabetes (Idd)10 locus on chromosome 3, originally identified by linkage analysis, was in fact due to three closely linked Idd loci: Idd10, Idd18.1, and Idd18.3. In this study, we define two additional Idd loci—Idd18.2 and Idd18.4—within the boundaries of this cluster of disease-associated genes. Idd18.2 is 1.31 Mb and contains 18 genes, including Ptpn22, which encodes a phosphatase that negatively regulates T and B cell signaling. The human ortholog of Ptpn22, PTPN22, is associated with numerous autoimmune diseases, including T1D. We, therefore, assessed Ptpn22 as a candidate for Idd18.2; resequencing of the NOD Ptpn22 allele revealed 183 single nucleotide polymorphisms with the C57BL/6J (B6) allele—6 exonic and 177 intronic. Functional studies showed higher expression of full-length Ptpn22 RNA and protein, and decreased TCR signaling in congenic strains with B6-derived Idd18.2 susceptibility alleles. The 953-kb Idd18.4 locus contains eight genes, including the candidate Cd2. The CD2 pathway is associated with the human autoimmune disease, multiple sclerosis, and mice with NOD-derived susceptibility alleles at Idd18.4 have lower CD2 expression on B cells. Furthermore, we observed that susceptibility alleles at Idd18.2 can mask the protection provided by Idd10/Cd101 or Idd18.1/Vav3 and Idd18.3. In summary, we describe two new T1D loci, Idd18.2 and Idd18.4, candidate genes within each region, and demonstrate the complex nature of genetic interactions underlying the development of T1D in the NOD mouse model