The role of HLA-DQ8 beta57 polymorphism in the anti-gluten T-cell response in coeliac disease.

Major histocompatibility complex (MHC) class II alleles HLA-DQ8 and the mouse homologue I-A(g7) lacking a canonical aspartic acid residue at position beta57 are associated with coeliac disease and type I diabetes. However, the role of this single polymorphism in disease initiation and progression remains poorly understood. The lack of Asp 57 creates a positively charged P9 pocket, which confers a preference for negatively charged peptides. Gluten lacks such peptides, but tissue transglutaminase (TG2) introduces negatively charged residues at defined positions into gluten T-cell epitopes by deamidating specific glutamine residues on the basis of their spacing to proline residues. The commonly accepted model, proposing that HLA-DQ8 simply favours binding of negatively charged peptides, does not take into account the fact that TG2 requires inflammation for activation and that T-cell responses against native gluten peptides are found, particularly in children. Here we show that beta57 polymorphism promotes the recruitment of T-cell receptors bearing a negative signature charge in the complementary determining region 3beta (CDR3beta) during the response against native gluten peptides presented by HLA-DQ8 in coeliac disease. These T cells showed a crossreactive and heteroclitic (stronger) response to deamidated gluten peptides. Furthermore, gluten peptide deamidation extended the T-cell-receptor repertoire by relieving the requirement for a charged residue in CDR3beta. Thus, the lack of a negative charge at position beta57 in MHC class II was met by negatively charged residues in the T-cell receptor or in the peptide, the combination of which might explain the role of HLA-DQ8 in amplifying the T-cell response against dietary gluten.

Effects of behavioral therapy or pharmacotherapy on brain glucose metabolism in subjects with obsessive-compulsive disorder as assessed by brain FDG PET.

This prospective study investigated the effect of pharmacotherapy (PT) and cognitive behavioral therapy (CBT) on cerebral glucose metabolism in adults with obsessive-compulsive disorder (OCD). Dynamic positron emission tomography (PET) of the brain with F-18-fluorodeoxyglucose (FDG) was performed before and after treatment in 16 subjects diagnosed for OCD for at least 2 years (PT: n=7). Pre-to-post-treatment change of scaled local metabolic rate of glucose (SLMRGlc) was assessed separately in therapy responders and non-responders. Correlation was tested between SLMRGlc change and change of OCD, depression, or anxiety symptoms. SLMRGlc increased in the right caudate after successful therapy. The increase tended to correlate with the improvement of OCD symptom severity. The finding of increased local caudate activity after successful therapy is in contrast to most previous studies. Possible explanations include effects of therapy on concomitant depression symptoms and/or the large proportion of early-onset OCD in the present sample.

Notch/Delta signaling constrains reengineering of pro-T cells by PU.1.

PU.1 is essential for early stages of mouse T cell development but antagonizes it if expressed constitutively. Two separable mechanisms are involved: attenuation and diversion. Dysregulated PU.1 expression inhibits pro-T cell survival, proliferation, and passage through beta-selection by blocking essential T cell transcription factors, signaling molecules, and Rag gene expression, which expression of a rearranged T cell antigen receptor transgene cannot rescue. However, Bcl2 transgenic cells are protected from this attenuation and may even undergo beta-selection, as shown by PU.1 transduction of defined subsets of Bcl2 transgenic fetal thymocytes with differentiation in OP9-DL1 and OP9 control cultures. The outcome of PU.1 expression in these cells depends on Notch/Delta signaling. PU.1 can efficiently divert thymocytes toward a myeloid-like state with multigene regulatory changes, but Notch/Delta signaling vetoes diversion. Gene expression analysis distinguishes sets of critical T lineage regulatory genes with different combinatorial responses to PU.1 and Notch/Delta signals, suggesting particular importance for inhibition of E proteins, Myb, and/or Gfi1 (growth factor independence 1) in diversion. However, Notch signaling only protects against diversion of cells that have undergone T lineage specification after Thy-1 and CD25 up-regulation. The results imply that in T cell precursors, Notch/Delta signaling normally acts to modulate and channel PU.1 transcriptional activities during the stages from T lineage specification until commitment.

Subversion of T lineage commitment by PU.1 in a clonal cell line system.

Specification of mammalian T lymphocytes involves prolonged developmental plasticity even after lineage-specific gene expression begins. Expression of transcription factor PU.1 may maintain some myeloid-like developmental alternatives until commitment. Commitment could reflect PU.1 shutoff, resistance to PU.1 effects, and/or imposition of a suicide penalty for diversion. Here, we describe subclones from the SCID.adh murine thymic lymphoma, adh.2C2 and adh.6D4, that represent a new tool for probing these mechanisms. PU.1 can induce many adh.2C2 cells to undergo diversion to a myeloid-like phenotype, in an all-or-none fashion with multiple, coordinate gene expression changes; adh.6D4 cells resist diversion, and most die. Diversion depends on the PU.1 Ets domain but not on known interactions in the PEST or Q-rich domains, although the Q-rich domain enhances diversion frequency. Protein kinase C/MAP kinase stimulation can make adh.6D4 cells permissive for diversion without protecting from suicide. These results show distinct roles for regulated cell death and another stimulation-sensitive function that establishes a threshold for diversion competence. PU.1 also diverts normal T-cell precursors from wild type or Bcl2-transgenic mice to a myeloid-like phenotype, upon transduction in short-term culture. The adh.2C2 and adh.6D4 clones thus provide an accessible system for defining mechanisms controlling developmental plasticity in early T-cell development.

Constitutive expression of PU.1 in fetal hematopoietic progenitors blocks T cell development at the pro-T cell stage.

The essential hematopoietic transcription factor PU.1 is expressed in multipotent thymic precursors but downregulated during T lineage commitment. The significance of PU.1 downregulation was tested using retroviral vectors to force hematopoietic precursors to maintain PU.1 expression during differentiation in fetal thymic organ culture. PU.1 reduced thymocyte expansion and blocked development at the pro-T cell stage. PU.1-expressing cells could be rescued by switching to conditions permissive for macrophage development; thus, the inhibition depends on both lineage and developmental stage. An intact DNA binding domain was required for these effects. PU.1 expression can downregulate pre-Talpha, Rag-1, and Rag-2 in a dose-dependent manner, and higher PU.1 levels induce Mac-1 and Id-2. Thus, downregulation of PU.1 is specifically required for progression in the T cell lineage.