DNA replication is undeniably important for life and as a consequence, cells have evolved mechanisms to monitor replication fidelity and to coordinate completion of replication with other cell cycle events. The goal of this project is to understand how cells choreograph the duplication of their chromosomes, and how defects in DNA replication may contribute to some recently discovered disorders in humans. Chromosome replication in eukaryotes is a process that involves the regulation of multiple initiation sites (origins) per chromosome. Origins show variation in timing-not all origins fire at the same time in S phase-and in efficiency-not all origins necessarily firing in every cell cycle. In addition, cells appea to maintain a backup set of origins (dormant origins) for use under special circumstances and these origins may differ in their basic control properties. Understanding how origin use is regulated is therefore critical for understanding how genome integrity is maintained. This notion is underscored by three recently described genetic disorders with links to replication defects: 1) Meier-Gorlin Syndrome with its point mutations in genes essential for origin licensing;2) a point mutation in a gene encoding part of the replicative helicase that is associated with breast cancer in mice and growth abnormalities in humans;and 3) human segmental triplications with an inverted central copy that can be explained by a replication fork error. Yeast is an ideal model organism for studying DNA replication because of its small chromosomes, well defined origin sequences, ease of altering chromosome structure, and exceptional systems for genetic and genomic analysis. This project will apply a combination of molecular, genetic and genomics approaches to: ? uncover modulators of origin efficiency and timing in yeast, to explore/investigate how orderly replication contributes to genome stability ? use point mutations in the yeast orthologs of genes involved in human disorders to understand the downstream consequences of defects in replication as they relate to human health and disease, and to identify genetic and chemical suppressors of the defects ? determine whether DNA replication errors are responsible for a particular class of inverted amplicons that can contribute to cancer progression and congenital copy number variants. This project will address outstanding questions regarding the biology of chromosome replication control: why do origins initiate replication at different times, what distinguishes origins in different temporal categories, what i the molecular basis for inefficient origins, and what defects in replication fork progression predispose chromosomal loci to inverted amplification? In addition, this work will give insight int human disorders resulting from errors in chromosome replication and could lead to diagnostic or therapeutic advances.

Public Health Relevance

Brewer Errors in chromosomal DNA duplication are a major source of genomic instability and are associated with a number of human disorders including cancer and some developmental disorders. We propose to continue our investigations into the mechanisms that modulate the orderly progression of chromosome replication and a direct role that replication origins may play in leading to genome instability. This work will expand our understanding of the mechanisms regulating chromosome duplication and illuminate questions of importance to human health.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM018926-42
Application #
8502226
Study Section
Molecular Genetics A Study Section (MGA)
Program Officer
Reddy, Michael K
Project Start
1976-01-01
Project End
2017-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
42
Fiscal Year
2013
Total Cost
$550,865
Indirect Cost
$187,754
Name
University of Washington
Department
Genetics
Type
Schools of Medicine
DUNS #
605799469
City
Seattle
State
WA
Country
United States
Zip Code
98195
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Di Rienzi, Sara C; Lindstrom, Kimberly C; Lancaster, Ragina et al. (2011) Genetic, genomic, and molecular tools for studying the protoploid yeast, L. waltii. Yeast 28:137-51
Raghuraman, M K; Brewer, Bonita J (2009) Molecular analysis of the replication program in unicellular model organisms. Chromosome Res :
Feng, Wenyi; Raghuraman, M K; Brewer, Bonita J (2007) Mapping yeast origins of replication via single-stranded DNA detection. Methods 41:151-7
Feng, Wenyi; Collingwood, David; Boeck, Max E et al. (2006) Genomic mapping of single-stranded DNA in hydroxyurea-challenged yeasts identifies origins of replication. Nat Cell Biol 8:148-55