Asthma epigneomics

2. Biological Background
Immunological Memory, is how a body is able to maintain a protective immune system, as it can respond to
a pathogen more rapidly and effectively if that pathogen has been encountered before.
Asthma, a disease caused by airway inflammation and is caused by excessive response to an allergen, such
as grass. The main molecular feature of asthma is the excessive differentiation of CD4 T cells. The goal of
this study was to develop methods to profile active and poised enhancers in naïve and memory CD4 T cells
from healthy individual and asthmatic patients. An enhancer, is a short region of DNA that is bound to
proteins and increases the likelihood that transcription of that particular gene will occur. This team generated
maps of H3K4me2 marked cis-regulatory regions and identified a large number of cell type specific and
disease specific enhancers.
http://www.nature.com/nri/journal/v10/n12/fig_tab/nri2870_F1.htmlhttp://www.nature.com/nri/journal/v8/n3/images/nri2262-f6.jpg

3. Article Background and Summary
• Its understood that a feature of asthma is the accumulation, differentiation,
and function of memory CD4+T cells that produce type 2 cytokines. The
goal of this study was to develop methods to profile the active and poised
enhancers in naive and memory CD4+T cells from healthy individuals and
asthmatic patients.
• In this article, the researchers isolated naïve and memory CD4 T cells from
peripheral blood of 24 subjects (12 healthy and 12 asthma). After isolation,
the CD4 memory T cells were divided by their surface expression of CCR4
(chemokine receptor). These cells were then sequenced for both RNA-seq
and ChIP-seq to identify the DNA regions associated with H3K4me2. A
total of 120 high-throughput whole genome ChIP-seq assays were
completed.

4. Data Information
• All done on AB 5500 Genetic Analyzer
• ChIP-Seq: 146 Hs biosamples, Single Ended GSE53646
• 619 ccr4+ t cells, 529 ccr4- t cells, 640 naïve cells runs
• ChIP-Seq: 26 Mm biosamples, Single Ended GSE53644
• 24 1k cells, 18 10k cells, 72 100k cells, 42 2m cells runs
• cell type:D10.G4.1 cells, ATCC
• chip antibody:H3K4me2 (clone Y47, lot#YH041501C;
Abcam)
• source name: mouse derived lymphoblastic cell line
(D10.G4.1 cells, ATCC)
• RNA-Seq: 49 Hs biosamples, Single Ended GSE55320
• Healthy patients: 88 ccr4+ t cells, 33 ccr4- t cells, 77
naïve t cells runs
• Asthmatic patients: 121 ccr4+ t cells, 99 ccr4- t cells runs,
121 naïve t cells runs
This study developed methods to profile the active and poised enhancers in naive and memory CD4+T cells.

5. T-Bioinfo Analysis Steps
Machine LearningRNA-seq Data Analysis
(mapping reads and generation of RSEM table)
ChIP-seq Data Analysis
(Mapping of reads and BS analysis)
Gene and Isoform expression profiles in
Th0, Th1, and Th2 cells of healthy
individual and asthma patients
H3K4me2 enrichment map for 500
bp-long in Th0, Th1, and Th2 cells of
healthy individual and asthma patients
Unsupervised BiAssociation and
clustering approach allowed
identification of some specific samples.
As well as Batch Effect Correction was
used in this project.

6. The basic strategy was to screen for genes whose
pattern of transcription and epigenetic modification
accounted for a particular T-cell phenotype and the
health-condition of the donor.
1) First, we determined components of PCA that reveal
biologically meaningful separation of groups
(Healthy/Unhealthy, Th1/Th2/Naive)
2) Second, we adopted factor regression analysis, a
procedure permitting investigation of relationships
between variables to identify those that produce
particular outcomes (in our case (Healthy/Unhealthy,
Th1/Th2/Naïve).
Both, the analysis of PCA components and factor
regression analysis helped to identify genes that
produce joined effects of cell type and health condition.
In other words it helped us to find genes and epigenetic
control regions, which while being distinctly regulated in
Naive, Th1 or Th2 cells simultaneously accounted for the
occurrence of asthma.