X-Ray Crystallography



Profile

Dr. Doublié received her Ph.D. in Biochemistry and Biophysics in 1993 under the direction of Charles W. Carter at the University of North Carolina at Chapel Hill. She did postdoctoral work with Stephen Cusack at the EMBL outstation in Grenoble and with Tom Ellenberger at Harvard Medical School where she solved the crystal structure of the ternary complex of T7 DNA polymerase. She joined the UVM faculty in October 1998.

Research Description

Modifications in DNA or RNA, as part of normal cellular processes or as aberrations, can have profound biological consequences. The major thrust of my research program is to study these nucleic acid modifications in the context of the enzymes and proteins that generate and recognize them.

DNA polymerases, which otherwise faithfully replicate DNA, stumble when they encounter oxidative DNA lesions. Polymerases either will be blocked at the site of lesion, or bypass it. The latter case, referred to as translesion synthesis, may initiate an oncogenic process if the wrong base is inserted opposite the lesion. Uncovering the fundamental mechanisms underpinning translesion synthesis is paramount to understand the initial events of mutagenesis. We aim to elucidate at the atomic level the factors that influence the interactions between a replicative polymerase and DNA lesions. Our goal is to answer the following questions: How does a DNA polymerase sense the presence of a DNA lesion? What role does sequence context play in translesion synthesis? What triggers the transfer of DNA to the editing site in the event of a base mispair ?

Several DNA repair mechanisms are in place to minimize damage in DNA before DNA polymerases replicate the genome. One of these processes is called Base Excision Repair (BER). The first step in BER is carried out by DNA glycosylases, so named because they hydrolyze the N-glycosidic bond between a damaged base and its deoxyribose, leaving an apurinic or apyrimidinic site in DNA. The goal of this project is to delineate the structural features of the DNA glycosylases that are involved in recognition of DNA base damage produced by ionizing radiation. In particular, we intend to address the question of how enzymes with a similar active site architecture are able to recognize vastly different substrates.

Messenger RNA (mRNA) also undergoes modifications, this time as an essential step of the normal cell program. During processing of the 3′-end of mRNA, a specific endonucleolytic cleavage event precedes the addition of a poly(A) tail. Such maturation of 3′-ends is a key regulatory step in the expression of many genes. The cleavage and polyadenylation reactions are carried out by a multicomponent machinery of remarkable complexity. Our goal is to understand how the different components of the machinery interact to cleave then polyadenylate messenger RNAs.

Highlighted Publications

Coseno ME, Martin G, Gilmartin G, Keller W and Doublié S. Crystal Structure of the 25 kDa Subunit of Human Cleavage Factor Im. Nucleic Acids Res 2008 Jun;36(10):3474-3483
View the structure at PDB

Imamura K., Wallace SS and Doublié S. Structural characterization of a viral NEIL1 ortholog unliganded and bound to abasic site-containing DNA. J Biol Chem. 2009 Jul 22. [Epub ahead of print]
View the structure at PDB.

Faucher F, Duclos S, Bandaru V, Wallace SS and Doublié S. Crystal structures of two archaeal 8-oxoguanine DNA glycosylases provide structural insight into guanine/8-oxoguanine distinction. Structure. 2009 May 13;17(5):703-12.
View the structure at PDB.

Faucher F, Robey-Bond SM, Wallace SS and Doublié S. Structural characterization of Clostridium acetobutylicum 8-oxoguanine DNA glycosylase in its apo form and in complex with 8-oxodeoxyguanosine. J Mol Biol. 2009 Apr 3;387(3):669-79. Epub 2009 Feb 9.
View the structure at PDB.

Aller P, Ye Y, Wallace SS, J Burrows C, Doublié S.Crystal Structure of a Replicative DNA Polymerase Bound to the Oxidized Guanine Lesion Guanidinohydantoin. Biochemistry. 2010 Feb 25. [Epub ahead of print]

Faucher F, Wallace SS, Doublié S. The C-terminal Lysine of Ogg2 DNA Glycosylases is a Major Molecular Determinant for Guanine/8-Oxoguanine Distinction. J Mol Biol. 2010 Mar 19;397(1):46-56.

Yang Q, Gilmartin GM, Doublié S. Structural basis of UGUA recognition by the Nudix protein CFI(m)25 and implications for a regulatory role in mRNA 3′ processing. Proc Natl Acad Sci U S A. 2010 Jun 1;107(22):10062-7.

View all Doublie publications here.

* indicates equal contribution

Recent Publications

Sjolund A, Nemec AA, Paquet N, Prakash A, Sung P, Doublié S, Sweasy JB (2014) A germline polymorphism of thymine DNA glycosylase induces genomic instability and cellular transformation. PLoS Genet 10(11): e1004753.

Yousefzadeh MJ, Wyatt DW, Takata K, Mu Y, Hensley SC, Tomida J, Bylund GO, Doublié S, Johansson E, Ramsden DA, McBride KM, Wood RD (2014) Mechanism of suppression of chromosomal instability by DNA polymerase POLQ. PLoS Genet 10(10): e1004654.

Doublié S, Zahn KE (2014) Structural insights into eukaryotic DNA replication. Front Microbiol 5: 444.

Towle-Weicksel JB, Dalal S, Sohl CD, Doublié S, Anderson KS, Sweasy JB (2014) Fluorescence resonance energy transfer studies of DNA polymerase β: the critical role of fingers domain movements and a novel non-covalent step during nucleotide selection. J Biol Chem 289(23): 16541-50.

Eckenroth BE, Fleming AM, Sweasy JB, Burrows CJ, Doublié S (2014) Crystal structure of DNA polymerase β with DNA containing the base lesion spiroiminodihydantoin in a templating position. Biochemistry 53(13): 2075-7.

Zahn KE, Doublié S (2014) Look Ma, no PCNA: how DNA polymerase ε synthesizes long stretches of DNA without a processivity factor. Nat Struct Mol Biol 21(1): 12-4.

Prakash A, Carroll BL, Sweasy JB, Wallace SS, Doublié S (2014) Genome and cancer single nucleotide polymorphisms of the human NEIL1 DNA glycosylase: activity, structure, and the effect of editing. DNA Repair (Amst) 14: 17-26.

View all Doublie publications here.

Selected Awards

Pew Scholar in the Biomedical Sciences The Pew Scholars Program in the Biomedical Sciences is designed to support young investigators of outstanding promise in the basic and clinical sciences relevant to the advancement of human health. The funding of the awards is provided by The Pew Charitable Trusts. The award is intended to provide assured support, during their earlier years, for junior members of the faculty as they establish their laboratories. It is hoped that the assurance provided through the Program will encourage successful applicants to be more venturesome in their research and future applications for support than would otherwise be likely.
(2000-2004)

J. Walter Juckett Award Each year, VCC\’s long-time partner—the Lake Champlain Cancer Research Organization (LCCRO)—generously makes funding available for cancer research and academic awards to UVM faculty in several categories. The J. Walter Juckett Scholar Award is one such award. Historically, this award has laid the foundation for larger, NIH-funded grants.
This two-year grant is awarded every other year to an investigator who has a demonstrated record of sustained productivity in cancer research.
The J. Walter Juckett Scholar Award provides $75,000 over a two-year period. The award is non-renewable, and may not be used to support tenure track faculty salary. (2001-2002)


Sylvie Doublie, Ph.D.

Sylvie
Doublie, Ph.D.

Professor
Department of Microbiology & Molecular Genetics

 

802-656-9531
Office: E314A Given
Lab: E314 Given

Lab Members

   Brittany Carroll, Research Technician
   Brian E. Eckenroth, Research Specialist
   Aishwarya Prakash, Postdoctoral Fellow
   Ryota Yokose, Undergraduate Student

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