Abstract

Hypochlorous acid (HOCl), the primary product of activated neutrophils at sites of inflammation, can damage both DNA and associated histone proteins. HOCl damage to cytosine bases in DNA generates the chlorination damage products 5-chlorocytosine (ClC) and 5-chlorouracil (ClU) as well as the oxidation damage products 5- hydroxycytosine (HOC) and 5-hydroxyuracil (HOU). Histone damage by HOCl can be measured by quantitation of 3-chlorotyrosine and 3,5-dichlorotyrosine, stable and unique markers of protein damage caused by HOCl. The studies presented here investigate how and where these damage products occur and what implications they might have in the correlation between chronic inflammation and carcinogenesis. The first study develops a strategy to place ClC into synthetic oligonucleotides providing a tool to study the biochemical and biophysical properties in DNA. This allowed further studies that show ClC could perturb epigenetic patterns in DNA by causing inappropriate methyltransferase-mediated cytosine methylation and by increasing the binding affinity of methyl-binding proteins involved in subsequent histone modification and chromosome condensation. In order for ClC to have the epigenetic effects proposed, it would have to be formed specifically in a CpG dinucleotide in duplex DNA. Utilizing mass tagging to show sequence specific chlorination by HOCl, the data provides direct evidence for the formation of ClC in a CpG dinucleotide. Emerging data shows the importance of histone modifications on gene transcription, replication, and repair. The histone proteins are in close proximity to DNA and have higher reactivity of protein side chains than DNA. 3-Chlorotyrosine and 3,5-dichlorotyrosine were measured as stable markers of histone damage by HOCl. The formation of 3-chlorotyrosine and 3,5-dichlorotyrosine in the core histone proteins, H2A, H2b, H3, and H4, upon reaction with HOCl was observed and quantified. The preferential chlorination of tyrosine residues within 4 residues of a lysine or histidine was also observed. The results of this series of studies provide a better understanding of HOCl damage products of DNA and histone proteins at sites of inflammation and shed light on the mechanisms by which inflammation contributes to carcinogenesis.

LLU Discipline

Biochemistry

Department

Biochemistry

School

Graduate Studies

First Advisor

Lawrence C. Sowers

Second Advisor

Jonathan W. Neidigh

Third Advisor

Ubaldo Soto-Wegner

Fourth Advisor

R. Bruce Wilcox

Fifth Advisor

Kangling Zhang

Degree Name

Doctor of Philosophy (Medical Science)

Degree Level

Ph.D.

Year Degree Awarded

2008

Date (Title Page)

5-2008

Language

English

Library of Congress/MESH Subject Headings

Inflammation -- immunology -- dissertations; Immunity, Cellular; Cell Aging -- drug effects; Hypochlorous Acid -- biosynthesis; Chromatin; Tyrosine; DNA Damage; Chromosomes; Oxidants -- physiology; Oxidative Stress -- physiology.

Type

Dissertation

Page Count

xiv; 179

Digital Format

PDF

Digital Publisher

Loma Linda University Libraries

Usage Rights

This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.

Collection

Loma Linda University Electronic Theses and Dissertations

Collection Website

http://scholarsrepository.llu.edu/etd/

Repository

Loma Linda University. Del E. Webb Memorial Library. University Archives

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