TRANSCRIPTOME ANALYSIS OF T CELLS IN CHROMOSOME 22Q11.2 DELETION SYNDROME

Background Phenotypic variations of chromosome 22q11.2 deletion syndrome (22qDS) have no clear explanation. T cell lymphopenia in chromosome 22q11.2 deletion syndrome (22qDS) is related to varying degrees of thymic hypoplasia and contributes to the phenotypic heterogeneity. No phenotype correlation with genotype or deletion size is known for lymphopenia. We hypothesized that the T-cell transcriptome is different in 22qDS compared to healthy children and that gene expression in T cells can differentiate patients with low T cells compared to normal T cells. Methods Peripheral blood was collected from a convenience sample of participants aged 5-8 years. Standard immune function testing was performed. RNA sequencing was completed on isolated T cells using Illumina’s TruSeq technology. Differential gene expression profiles (q<0.05) of T cells between 22qDS and healthy controls were determined with Tuxedo Suite & String Tie pipelines. Bioinformatic analyses were implemented via Ingenuity Pathway Analysis and KEGG to identify enriched pathways. The Spearman correlation between gene expression and clinical variables were calculated using SAS (9.4, Cary, NC) (p-value<0.05). Results A total of 360 genes were differentially expressed between T cells of 22qDS patients (n=13) and healthy controls (n=6) (Log 2 fold change range (-2.0747, 15.6724)).When these 360 genes were tested for pathway enrichment, the top 5 pathways in T lymphocytes based on their p value included communication between innate and adaptive immune cells, cross talk between dendritic cells and natural killer cells, allograft rejection signaling, dendritic cell maturation, and B cell receptor signaling. The top 10 biological processes with differential expression included 36 immune response, 31 inflammatory response, 33 apoptotic process, 12 interferon gamma mediated signaling pathway, 14 nucleosome assembly, 16 defense response to virus, 8 lipopolysaccharide mediated signaling pathway, 7 positive regulation of NF-kappa B import into nucleus, 10 type I interferon signaling pathway, and 10 neutrophil chemotaxis genes. We compared gene expression between 22qDS participants with low T cell counts (n=7) and 22qDS participants with normal T cell counts (n=6) and found 94 genes that were differentially expressed (q<0.05) (Log2 fold change range (-4.5445, 5.1297)). We found 29 genes that correlated with T cell counts and subsets (CD3, CD4, CD8, and naïve helper T cells) (R≥0.8). Conclusions T-cell gene expression contributes to phenotypic heterogeneity in chromosome 22q11.2 deletion syndrome. T cell gene expression is different in 22qDS with low T cells compared to normal T cells. Differential gene expression can be used in future to develop biomarkers to differentiate between different phenotypes of 22qDS. Further, therapies can be personalized based on the phenotype using RNA therapeutics

Utility of next generation sequencing in clinical primary immunodeficiencies.

Primary immunodeficiencies (PIDs) are a group of genetically heterogeneous disorders that present with very similar symptoms, complicating definitive diagnosis. More than 240 genes have hitherto been associated with PIDs, of which more than 30 have been identified in the last 3 years. Next generation sequencing (NGS) of genomes or exomes of informative families has played a central role in the discovery of novel PID genes. Furthermore, NGS has the potential to transform clinical molecular testing for established PIDs, allowing all PID differential diagnoses to be tested at once, leading to increased diagnostic yield, while decreasing both the time and cost of obtaining a molecular diagnosis. Given that treatment of PID varies by disease gene, early achievement of a molecular diagnosis is likely to enhance treatment decisions and improve patient outcomes

Additional file 3: of Novel heterozygous pathogenic variants in CHUK in a patient with AEC-like phenotype, immune deficiencies and 1q21.1 microdeletion syndrome: a case report

Figure S3. Growth chart. A) Weight for age (kg). B) Length for age (cm). (TIFF 602 kb

Additional file 1: of Novel heterozygous pathogenic variants in CHUK in a patient with AEC-like phenotype, immune deficiencies and 1q21.1 microdeletion syndrome: a case report

Figure S1. DNA alignment of NGS data using Integrative Genomics Viewer (IGV). IGV snapshot of exon 13 of CHUK gene (NM_001278.3), located on chromosome 10, showing that the two variants (c.1365del, p.Arg457Aspfs*6; c.1388C > A, p.Thr463Lys) are present on different reads, indicating that they occurred on different chromosome (in trans). (TIFF 307 kb

Additional file 2: of Novel heterozygous pathogenic variants in CHUK in a patient with AEC-like phenotype, immune deficiencies and 1q21.1 microdeletion syndrome: a case report

Figure S2. The protein is composed of a protein kinase domain (orange), a leucine zipper region (green) and the NEMO binding-region (blue). Missense (gray) and loss of function (red) pathogenic variants found in the CHUK gene. Reported in this study (Bold). (TIFF 128 kb