Research
Research overview
Many, if not all cancers have unstable genomes. Genomic instability often arises from defects in DNA repair that can drive cancer development. Yet, the inability of cancer cells to faithfully repair DNA makes them vulnerable to DNA damaging cancer therapies, which are commonly used as first-line treatments.
The 3D architecture of the genome during DNA repair influences the balance between proper repair, which is a tumor suppressor mechanism, and pathological repair, which can cause oncogenic chromosome rearrangements. Defining the spatial organization of the genome during DNA repair following DNA damage will help develop a better understanding of cancer etiology and therapy mechanisms.
The overarching research goal of the Gautier Lab is to define how DNA lesions (e.g., DNA double-strand breaks and DNA interstrand crosslinks) and their repairs create genetic mutations and chromosome rearrangements within their various chromatin contexts and at specific cell cycle phases. To achieve this goal, the lab uses diverse experimental approaches, encompassing biochemistry, proteomics, live-cell imaging, super-resolution microscopy, Hi-C and genome-wide translocation sequencing. Experimental systems used in the lab include cell-free extracts that are used as a simple model system to study processes that govern genome stability, including DNA replication control, DNA repair and the cellular response to DNA damage, as well as cultured normal and tumor cells that are used to model and analyze biological responses to DNA damage.
Actin and actin-related proteins in 3D genomic reorganization following DNA damage
DNA repair is enabled within nuclear compartments or repair domains. Spatial organization of repair facilitates biochemical reactions and restricts reactions to specialized compartments to facilitate repair. Bringing DNA lesions into proximity also favors the creation of rare, aberrant genome rearrangements.
The Gautier Lab made the seminal discovery that mechanical forces drive the 3D arrangement of the genome following damage. The lab is now working to better define the mechanisms underpinning the assembly of DNA repair domains and to evaluate the physio-pathological consequences of assembling these domains following treatment with chemotherapeutic drugs.
Relevant publications:
Schrank BR, Aparicio T, Li Y, Chang W, Chait BT, Gundersen GG, Gottesman ME, Gautier J. Nature. 2018 Jul;559(7712):61-66. PMID: 29925947
Zagelbaum J, Schooley A, Zhao J, Schrank BR, Callen E, Zha S, Gottesman ME, Nussenzweig A, Rabadan R, Dekker J, Gautier J. Nat Struct Mol Biol. 2023 Jan;30(1):99-106. PMID: 36564591
Mechanisms of genomic rearrangements at sites of DNA double-strand breaks
Insertions and deletions (indels) of one or more bases and translocations are common aberrations in cancer genomes. These rearrangements result from various cellular mechanisms, often including DNA replication and transcription, leading to pathological DNA repair. When rearrangements form within DNA coding sequences, frameshift mutations can occur, which may create a protein with reduced or altered function. When rearrangements occur in non-coding regions, they can alter gene expression or splicing patterns.
The Gautier Lab developed a new assay, Indel-seq, to assess indels resulting from both precise experimental introduction and spontaneous genome instability. They determined that templated insertions use templates from across the genome, and that such insertions require contact between the donor and acceptor loci, followed by homologous recombination. Their data are consistent with a model in which the broken acceptor site either anneals to a resected DNA break or invades the displaced strand of a transcription bubble or R-loop, followed by DNA synthesis displacement and ligation via non-homologous end joining. (See model to the left.)
Relevant publications:
Min J, Zhao J, Zagelbaum J, Lee J, Takahashi S, Cummings P, Schooley A, Dekker J, Gottesman ME, Rabadan R, Gautier J. Mol Cell. 2023 Jul 20;83(14):2434-2448.e7. PMID: 37402370
Zagelbaum J, Schooley A, Zhao J, Schrank BR, Callen E, Zha S, Gottesman ME, Nussenzweig A, Rabadan R, Dekker J, Gautier J. Nat Struct Mol Biol. 2023 Jan;30(1):99-106. PMID: 36564591
Transcription-coupled DNA repair and transcription-driven genome instability
In cells, ongoing transcription continuously scans the genome, detects DNA lesions and facilitates their repair, making transcription a tumor suppressor mechanism. However, transcription can also promote genome instability by exposing fragile intermediates, such as R-loops, to aberrant genetic alterations.
DNA interstrand crosslinks (ICLs, see depiction on the left) form due to exposure to endogenous aldehyde mutagens as well as from the action of chemotherapeutic drugs (e.g. mitomycin C, cisplatin and psoralens). An ICL is a highly toxic, covalent bond that binds two complementary DNA strands together, preventing their separation. This creates a roadblock to DNA replication, a prominent sensor of ICL lesions. The Gautier Lab demonstrated that ICLs could be sensed and repaired outside of S-phase via multiple mechanisms, including mismatch repair (MMR).
Relevant publications:
Ben-Yehoyada M, Wang LC, Kozekov ID, Rizzo CJ, Gottesman ME, Gautier J. Mol Cell. 2009 Sep 11;35(5):704-15. PMID: 19748363.
Williams HL, Gottesman ME, Gautier J. Mol Cell. 2012 Jul 13;47(1):140-7. PMID: 22658724
Kato N, Kawasoe Y, Williams H, Coates E, Roy U, Shi Y, Beese LS, Sch?rer OD, Yan H, Gottesman ME, Takahashi TS, Gautier J. Cell Rep. 2017 Oct 31;21(5):1375-1385. PMID: 29091773
Recently, the Gautier Lab discovered that ultraviolet-stimulated scaffold protein A (UVSSA) is required for transcription-coupled ICL repair and that inactivation of UVSSA not only sensitizes cells to ICL-inducing drugs but also impedes the ICL repair pathway. In addition, they determined that UVSSA interacts with transcribing RNA polymerase (Pol II) and transcription-coupled DNA repair complex factors including CSB, CSA, and transcription factor IIH (TFIIH). The lab is now working to better understand the mechanisms of transcription-coupled interstrand crosslink repair, as well as better defining the role of UVSSA in the cell.
Notably, the Gautier Lab also discovered that UVSSA helps protect cells from the transcriptional stress caused by MYC overexpression, which is common in many cancers. This guardian role of UVSSA relies on its ability to interact with RNA polymerase (Pol II).
Relevant publications:
Liebau RC, Waters C, Ahmed A, Soni RK, Gautier J. DNA Repair (Amst). 2024 Nov;143:103771. PMID: 39383571
Sato M, Liebau RC, Liu Z, Liu L, Rabadan R, Gautier J. J Cell Biol. 2021 Feb 1;220(2):e201807163. PMID: 33404608