title	author	date	theme	aspectratio	fontsize
Bioinformatic analysis of complex, high-throughput genomic and epigenomic data in the context of $\mathsf{CD4}^{+}$ T-cell differentiation and diagnosis and treatment of transplant rejection	Ryan C. Thompson \ Su Lab \ The Scripps Research Institute	October 24, 2019	Boadilla	169	14pt

Organ transplants are a life-saving treatment

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36,528 transplants performed in the USA in 2018¹
100 transplants every day!
Over 113,000 people on the national transplant waiting list as of July 2019

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Organ donation statistics for the USA in 2018¹

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Types of grafts

A graft is categorized based on the relationship between donor and recipient:

. . .

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Autograft: Donor and recipient are the same individual
Allograft: Donor and recipient are different individuals of the same species
Xenograft: Donor and recipient are different species

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Recipient T-cells reject allogenic MHCs

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TCR binds to both antigen and MHC surface \vspace{10pt}
HLA genes encoding MHC proteins are highly polymorphic \vspace{10pt}
Variants in donor MHC can trigger the same T-cell response as a foreign antigen

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$TCR binding to self (right) and allogenic (left) MHC\footnotemark$ { height=70% } :::

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\footnotetext[3]{\href{https://doi.org/10.1016/j.cell.2007.01.048}{Colf, Bankovich, et al. "How a Single T Cell Receptor Recognizes Both Self and Foreign MHC". In: Cell (2007)}}

Allograft rejection is a major long-term problem

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Rejection is treated with immune suppressive drugs

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Graft recipient must take immune suppressive drugs indefinitely
Graft is monitored for rejection and dosage adjusted over time
Immune suppression is a delicate balance: too much and too little are both problematic.

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Memory cells: faster, stronger, and more independent

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Compared to naïve cells, memory cells:

respond to a lower antigen concentration
respond more strongly at any given antigen concentration
require less co-stimulation
are somewhat independent of some types of co-stimulation required by naïve cells
evolve over time to respond even more strongly to their antigen

Result:

Memory cells require progressively higher doses of immune suppresive drugs
Dosage cannot be increased indefinitely without compromising the immune system's ability to fight infection

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3 problems relating to transplant rejection

1. How are memory cells different from naïve?

\onslide<2->{Genome-wide epigenetic analysis of H3K4 and H3K27 methylation in naïve and memory $\mathsf{CD4}^{+}$ T-cell activation}

2. How can we diagnose rejection noninvasively?

\onslide<3->{Improving array-based diagnostics for transplant rejection by optimizing data preprocessing}

3. How can we evaluate effects of a rejection treatment?

\onslide<4->{Globin-blocking for more effective blood RNA-seq analysis in primate animal model for experimental graft rejection treatment}

Today's focus

\Large 1. How are memory cells different from naïve?

\Large

Genome-wide epigenetic analysis of H3K4 and H3K27 methylation in naïve and memory $\mathsf{CD4}^{+}$ T-cell activation

We need a better understanding of immune memory

Cell surface markers fairly well-characterized
But internal mechanisms poorly understood

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\large

Hypothesis: Epigenetic regulation of gene expression through histone modification is involved in $\mathsf{CD4}^{+}$ T-cell activation and memory.

Which histone marks are we looking at?

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H3K4me3: "activating" mark associated with active transcription
H3K4me2: Correlated with H3K4me3, hypothesized "poised" state
H3K27me3: "repressive" mark associated with inactive genes

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. . .

\vfill

All involved in T-cell differentiation, but activation dynamics unexplored

ChIP-seq measures DNA bound to marked histones³

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Experimental design

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\footnotesize

Data generated by Sarah Lamere, published in GEO as GSE73214

Time points capture phases of immune response

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A few intermediate analysis steps are required

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Questions to focus on

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How do we define the "promoter region" for each gene? \vspace{10pt}
How do these histone marks behave in promoter regions? \vspace{10pt}
What can these histone marks tell us about T-cell activation and differentiation?

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First question

\centering \LARGE

How do we define the "promoter region" for each gene?

Histone modifications occur on consecutive histones

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Histone modifications occur on consecutive histones

\begin{figure} \centering \only<1>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/CCF-plots-A-SVG.png}} \only<2>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/CCF-plots-B-SVG.png}} \only<3>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/CCF-plots-C-SVG.png}} \caption{Strand cross-correlation plots show histone-sized wave pattern} \end{figure}

SICER identifies enriched regions across the genome

IDR identifies reproducible enriched regions

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Finding enriched regions across the genome

Each histone mark has an "effective promoter radius"

Peaks in promoters correlate with gene expression

\begin{figure} \centering \only<1>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/FPKM-by-Peak-Violin-Plots-A-SVG.png}} \only<2>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/FPKM-by-Peak-Violin-Plots-B-SVG.png}} \only<3>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/FPKM-by-Peak-Violin-Plots-C-SVG.png}} \only<4>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/FPKM-by-Peak-Violin-Plots-D-SVG.png}} \only<5>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/FPKM-by-Peak-Violin-Plots-Z-SVG.png}} \caption{Expression distributions of genes with and without promoter peaks} \end{figure}

First question

\centering \LARGE

How do we define the "promoter region" for each gene?

Answer: Define the promoter region empirically!

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H3K4me2, H3K4me3, and H3K27me3 occur in broad regions across the genome
Enriched regions occur more commonly near promoters
Each histone mark has its own "effective promoter radius"
Presence or absence of a peak within this radius is correlated with gene expression

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::: {.column width="50%"} \centering \only<1>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/CCF-plots-A-SVG.png}} \only<2>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/Promoter-Peak-Distance-Profile-SVG.pdf}} \only<3>{\includegraphics[width=\textwidth,height=0.7\textheight]{graphics/presentation/FPKM-by-Peak-Violin-Plots-A-SVG.png}} ::: ::::::::::

Next question

\centering \LARGE

How do these histone marks behave in promoter regions?

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Does the position of a histone modification within a gene promoter matter to that gene's expression, or is it merely the presence or absence anywhere within the promoter?

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H3K4me2 promoter neighborhood K-means clusters

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