Deciphering Networks Controlling DNA Amplification

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Principal Investigator: Johnathan R. Whetstine
Organization: RESEARCH INST OF FOX CHASE CAN CTR
Fiscal Year: 2024
Award: $467,500
Funding agency: National Institute of General Medical Sciences

PROJECT SUMMARY
DNA amplification is associated with pathological states such as neurological disorders, cardiac disease and
cancer. At least 50% of the amplifications in cancer are transient extrachromosomal DNA (ecDNA). The transient
behavior contributes to copy number plasticity, results in heterogeneous oncogene expression and alters
therapeutic response. The unanswered question remains as to whether distinct mechanisms control ecDNA copy
gains within cells and how they impact copy gain events associated with disease. My overall goal is to define the
principles regulating selective DNA copy gains and the associated plasticity, so we may control these events.
My laboratory was the first to discover a molecular basis for extrachromosomal transient site-specific DNA copy
gains (TSSGs) in the human genome. Specifically, we identified the first enzyme capable of driving site-specific
ecDNA amplification [the histone 3 lysine 9 and 36 tri-demethylase (H3K9/36me3) KDM4A] and demonstrated a
fundamental role for epigenetic states in controlling the predilection of specific DNA regions to rereplicate and
amplify. We discovered seven more chromatin enzymes- lysine methyltransferases (KMTs) and demethylases
(KDMs)- that function in concert to control site-specific amplification in both non-cancer and cancer cells. These
studies established a critical role for chromatin factors and their associated states in regulating DNA
amplifications. With this NIGMS R35, my laboratory will expand our studies in order to elucidate: 1) the
fundamental mechanisms controlling DNA amplification; 2) the molecular processes and characteristics
promoting or preventing DNA amplification; and 3) the relationship between ecDNA generation and the
associated RNA heterogeneity/DNA mutation burden. We will address these points by leveraging microscopy-
based screens using genetic and chemical tools in order to identify key amplifiers, and in turn, generate
epigenome profiles coupled to genome organization maps associated with these pathways so that molecular
features controlling DNA amplification are resolved. These studies will also be coupled to state-of-the-art long
read sequencing and single cell (DNA and RNA) sequencing strategies so that the associated heterogeneity
within the cell population and individual cells can be correlated with the effect of the amplifier on TSSGs. These
studies are being conducted in non-transformed cells that have a nearly diploid genome so that additional
genomic anomalies and mutations do not impact these studies. Collectively, the data generated from these
studies will increase our knowledge about the molecular features governing DNA copy gains and the associated
heterogeneity, which will resolve novel biomarkers and therapeutic targets in order to control copy number-
associated diseases in the years ahead.

Terms: <Amplifiers><Automobile Driving><Behavior><Cancer Genes><Cancer-Promoting Gene><Cancers><Cardiac Diseases><Cardiac Disorders><Cell Body><Cells><Characteristics><Chemicals><Chromatin><Coupled><DNA><DNA Alteration><DNA Sequence Alteration><DNA amplification><DNA mutation><Data><Deoxyribonucleic Acid><Diploid><Diploidy><Disease><Disorder><EC 2.1.1><Enzyme Gene><Enzymes><Epigenetic><Epigenetic Change><Epigenetic Mechanism><Epigenetic Process><Event><Gene Amplification><Generations><Genetic><Genetic Alteration><Genetic Change><Genetic defect><Genetic mutation><Genome><Genomics><Goals><Heart Diseases><Heterogeneity><Histones><Human Genome><Individual><Knowledge><L-Lysine><Laboratories><Lysine><Malignant Cell><Malignant Neoplasms><Malignant Tumor><Maps><Methyltransferase><Microscopy><Molecular><Mutation><NIGMS><National Institute of General Medical Sciences><Nervous System Diseases><Nervous System Disorder><Neurologic Disorders><Neurological Disorders><Non-Polyadenylated RNA><Nuclear><Oncogenes><Pathologic><Pathway interactions><Population><Process><RNA><RNA Gene Products><Ribonucleic Acid><Role><Sequence Alteration><Site><Transforming Genes><cancer cell><driving><epigenetically><epigenome><extrachromosomal DNA><genome mutation><genomic alteration><heart disorder><human whole genome><insight><malignancy><methylase><natural gene amplification><neoplasm/cancer><neurological disease><new drug target><new druggable target><new marker><new pharmacotherapy target><new therapeutic target><new therapy target><novel biomarker><novel drug target><novel druggable target><novel marker><novel pharmacotherapy target><novel therapeutic target><novel therapy target><pathway><prevent><preventing><response to therapy><response to treatment><social role><therapeutic response><therapeutic target><therapy response><tool><transmethylase><treatment response><treatment responsiveness>