Dr. Sylvie Doublie
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.
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]
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.
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.
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.
Hyde SJ, Eckenroth BE, Smith BA, Eberley WA, Heintz NH, Jackman JE, Doublié S. tRNAHis guanylyltransferase (THG1), a unique 3′-5′ nucleotidyl transferase, shares unexpected structural homology with canonical 5′-3′ DNA polymerases. Proc Natl Acad Sci U S A. 2010 Nov 8. [Epub ahead of print]
Yang Q, Gilmartin GM, Doublié S. The structure of human Cleavage Factor Im hints at functions beyond UGUA-specific RNA binding: A role in alternative polyadenylation and a potential link to 5′ capping and splicing. RNA Biol. 2011 Sep 1;8(5). [Epub ahead of print]
Yang Q, Doublié S. Structural biology of poly(A) site definition. Wiley Interdiscip Rev RNA. 2011 Sep;2(5):732-47.
Zahn KE, Tchesnokov EP, Götte M, Doublié S. Phosphonoformic acid inhibits viral replication by trapping the closed form of the DNA polymerase. J Biol Chem. 2011 Jul 15;286(28):25246-55.
Aller P, Duclos S, Wallace SS, Doublié S. A crystallographic study of the role of sequence context in thymine glycol bypass by a replicative DNA polymerase serendipitously sheds light on the exonuclease complex. J Mol Biol. 2011 Sep 9;412(1):22-34.
Yang Q, Coseno M, Gilmartin GM, Doublié S. Crystal structure of a human cleavage factor CFI(m)25/CFI(m)68/RNA complex provides an insight into poly(A) site recognition and RNA looping. Structure. 2011 Mar 9;19(3):368-77.
Zahn KE, Wallace SS, Doublié S. DNA polymerases provide a canon of strategies for translesion synthesis past oxidatively generated lesions. Curr Opin Struct Biol. 2011 Jun;21(3):358-69.