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Series GSE134883 Query DataSets for GSE134883
Status Public on Jul 25, 2019
Title The Position Beta57 of IAg7 Controls the Early Anti-Insulin Response and Onset of Diabetes in NOD mice to Link MHC and Disease [sequencing]
Organism Mus musculus
Experiment type Expression profiling by high throughput sequencing
Summary Abstract- The class II region of the Major Histocompatibility locus is the main contributor to the genetic susceptibility to type 1 diabetes (T1D). The loss of an aspartic acid at position 57 of diabetogenic HLA-DQ8 chains supports this association; it influences the recognition of peptides in the context of HLA-DQ8, and I-Ag7 using a mechanism termed the P9 switch. Here, we built register-specific insulin MHC tetramers, Ins12-20 and Ins13-21, to examine anti-insulin CD4 T cell responses during the early pre-diabetic phase of disease in mice. A single cell gene expression analysis and single cell T cell receptor (TCR) sequencing of anti-insulin CD4 T cells performed in 6 and 12-week-old NOD mice, revealed tissue-specific gene expression signatures. TCR signaling and clonal expansion were found only in the islets of Langerhans. and produced either classical Th1 differentiation or an unusual Treg phenotype, independent of TCR usage. The early phase of the anti-insulin response was dominated by cells specific for Ins12-20, the register that supports a P9 switch mode of recognition. The presence of the switch was demonstrated by TCR sequencing, re-expression, mutagenesis, and functional testing of alpha/beta TCR pairs in vitro. The genetic correction of the beta57 mutation resulted in the disappearance of D/E residues in the CDR3beta of anti-Ins12-20 T cells, and inability of cells normally activated by a switch to recognize the Ins9-23 peptide. These results provide the first molecular mechanistic explanation that links the unique MHC class II polymorphism of T1D with the recognition of islet antigens and disease onset.
Overall design Murine CD4+ T cells, isolated from different tissues, were stained with IAg7 tetramers labelled with the fluorescent moiety PE. Cells stained with either tetramer were single cell sorted into single wells of 96 well plates. Single cell cDNA libraries were made according to the Fluidigm BioMark "Two Step Single Cell Gene Expression Using EvaGreen" protocol with primers for 96 genes as well as unique TCRalpha and TCRbeta chains. Next, TCR alpha and beta chains were amplified and each single well/cell was barcoded on both alpha and beta chains using the Fluidigm AccessArray. (The protocol, and the barcodes utilized, are published in detail in Holt et al; PMID 29224077). Library preparation was done by the TSRI Genomics Core and run on an Illumina MiSeq. Post-alignment, reads were run through the international ImmunoGenetics information system. Confirmed genomically encoded regions as well as CDR3 regions were analyzed via a custom R script.
Contributor(s) Gioia L, Holt M, Costanzo A, Sharma S, Abe B, Kain L, Nakayama M, Wan X, Su A, Mathews C, Chen YG, Unanue E, Teyton L
Citation(s) 31471352
Submission date Jul 25, 2019
Last update date Oct 24, 2019
Contact name Luc Teyton
Organization name TSRI
Lab Teyton Lab
Street address 10550 North Torrey Pines Road
City La Jolla
State/province CA
ZIP/Postal code 92037
Country USA
Platforms (1)
GPL16417 Illumina MiSeq (Mus musculus)
Samples (16)
GSM3976424 M142-101515
GSM3976425 M230-07-06-16
GSM3976426 M225-06-24-16
This SubSeries is part of SuperSeries:
GSE134885 The Position Beta57 of IAg7 Controls the Early Anti-Insulin Response and Onset of Diabetes in NOD mice to Link MHC and Disease
BioProject PRJNA556687
SRA SRP216405

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Supplementary file Size Download File type/resource
GSE134883_ALL_T1D_TCRA_TCRB_Master_List_Ms.xlsx 267.4 Kb (ftp)(http) XLSX
GSE134883_RAW.tar 800.0 Kb (http)(custom) TAR (of XLS, XLSX)
GSE134883_Sample_metadata.xlsx 82.2 Kb (ftp)(http) XLSX
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Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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