Ying Liu – University of Copenhagen

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Ying Liu

The general research interest of Liu lab is to understand the various mechanisms that cause genome instability in human cells, how cells counteract these threats, and why some regions in the genome are unusually susceptible to instability (e.g. common or rare fragile sites). In CCS, Liu group has the following three research focuses:

1) The characterization of DNA/RNA hybrids (R-loops) associated with common fragile sites

Common fragile sites (CFSs) are hot spots for deletions and chromosome rearrangements in cancer [1]. It is well established that incomplete replication can cause a delay in chromatin condensation that leads to the ‘expression’ of CFSs (detected as breaks or gaps on metaphase chromosomes) [2, 3]. CFS regions have some intrinsic characteristics that predispose them to ‘fragility’ under replication stress conditions (RS). These include, 1) containing repeat sequences that could form secondary structures (e.g. hairpins or G-quadruplexes); 2) lack of replication origins; and 3) containing long genes or highly transcribed genes where replication forks are prone to collide with the messenger RNAs (e.g. regions where R-loops can form) [2, 4-8]. We have previously discovered that a process called mitotic DNA synthesis (MiDAS) that operates in mitosis is a strategy used by human cells to rescue the incomplete replication at those loci, particularly in cancers cells [9-12]. Our current focus is to identify the mechanisms employed by the cells to resolve stalled replication forks in the regions prone to form R-loops (Figure 1).

A diagram illustration of a head-on collision between a replication and a transcription where a R-loop has formed
Figure 1, A diagram illustration of a head-on collision between a replication and a transcription where a R-loop has formed

2) The analysis of folate deficiency induced genome instability

Folate deficiency is known to be associated with a diverse range of human disorders including fetal neural tube defects, age-associated dementia, infertility, and some type of cancers. Intriguingly, folate deficiency is known can cause the expression of group of rare fragile sites, all of which contain long stretch of CGG simple repeats. The most well studied locus of this kind is called FRAXA that is associated with Fragile X syndrome (FXS). We use FXS cells as a model to study how the FRAXA locus is affected by folate deprivation. Our data suggest that folate deprivation triggers extensive missegregation of this locus during cell division. Moreover, the entire chromosome X becomes unstable during a period of long-term folate deprivation [13]. We are currently working on, 1) the characterization of the pathways employed by the cells to repair the collapsed replication folks caused by folate deficiency; and 2) the identification of genomic regions that are most vulnerable to folate deficiency (Figure 2).

Figure 2, A diagram illustration of the consequence of folate deficiency in cells with a abnormally long CGGs at FRAXA.

Figure 2, A diagram illustration of the consequence of folate deficiency in cells with a abnormally long CGGs at FRAXA. In S phase, the replication of long CGGs at FRAXA is perturbed due to folate deficiency. This leads to a greater chance of collapse of replication forks within FRAXA. The fork breakage initiates homologous recombination (HR) repair in late S phase. A proportion of the HR intermediates (Holliday junctions, HJ; or D-loops) fail to be resolved prior to anaphase, leading to the formation of ssDNA bridges bound by RPA.  The FRAXA-containing bridges are ultimately either lost or retained in micronuclei in the next generation of G1 cells. Unresolved bridges might also trigger cytokinesis failure and hence promote the generation of binucleated cells, which tend to generate aneuploid progeny.

 3) Post-translational modification of proteins by sumoylation in response to DNA replication stress

In response to replication-associated DNA damage, human cells activate a highly conserved signalling network to prevent irreversible breakdown of DNA replication forks. One component of this DNA damage response is post-translational modification (i.e. phosphorylation, acetylation, mono- or poly-ubiquitylation and SUMOylation). While much is known about how phosphorylation affects protein function, little information is available on the SUMOylation of proteins following cellular stress. From an unbiased mass spectrometry based proteomic study, we have previously shown that, in response to RS (a condition that could initiate CFS expression), SUMO2 was conjugated to a panel of proteins in S phase in human cells [14, 15]. Interestingly, POLD3, a subunit of human polymerase delta, was found to be modified most significantly amongst these proteins [14]. We are currently carrying out research to characterise how POLD3 SUMOylation by SUMO2 might play a role in the cellular response to RS.

 References:

  1. R.I. Richards, Fragile and unstable chromosomes in cancer: causes and consequences, Trends in genetics : TIG, 17 (2001) 339-345.
  2. A. Helmrich, M. Ballarino, L. Tora, Collisions between Replication and Transcription Complexes Cause Common Fragile Site Instability at the Longest Human Genes, Molecular Cell, 44 (2011) 966-977.
  3. A. Letessier, G.A. Millot, S. Koundrioukoff, A.M. Lachages, N. Vogt, R.S. Hansen, B. Malfoy, O. Brison, M. Debatisse, Cell-type-specific replication initiation programs set fragility of the FRA3B fragile site, Nature, 470 (2011) 120-123.
  4. A.V. Barros, M.A. Wolski, V. Nogaroto, M.C. Almeida, O. Moreira-Filho, M.R. Vicari, Fragile sites, dysfunctional telomere and chromosome fusions: What is 5S rDNA role?, Gene, 608 (2017) 20-27.
  5. S.A. Hosseini, S. Horton, J.C. Saldivar, S. Miuma, M.R. Stampfer, N.A. Heerema, K. Huebner, Common Chromosome Fragile Sites in Human and Murine Epithelial Cells and FHIT/FRA3B Loss-Induced Global Genome Instability, Gene Chromosome Canc, 52 (2013) 1017-1029.
  6. C.J. McNees, A.M. Tejera, P. Martinez, M. Murga, F. Mulero, O. Fernandez-Capetillo, M.A. Blasco, ATR suppresses telomere fragility and recombination but is dispensable for elongation of short telomeres by telomerase, J Cell Biol, 188 (2010) 639-652.
  7. A. Sfeir, S.T. Kosiyatrakul, D. Hockemeyer, S.L. MacRae, J. Karlseder, C.L. Schildkraut, T. de Lange, Mammalian Telomeres Resemble Fragile Sites and Require TRF1 for Efficient Replication, Cell, 138 (2009) 90-103.
  8. I. Simonic, G.S. Gericke, The enigma of common fragile sites, Human Genetics, 97 (1996) 524-531.
  9. V.A. Bjerregaard, O. Ozer, I.D. Hickson, Y. Liu, The Detection and Analysis of Chromosome Fragile Sites, Methods Mol Biol, 1672 (2018) 471-482.
  10. L. Garribba, W. Wu, O. Ozer, R. Bhowmick, I.D. Hickson, Y. Liu, Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci, Method Enzymol, 601 (2018) 45-58.
  11. S. Minocherhomji, S. Ying, V.A. Bjerregaard, S. Bursomanno, A. Aleliunaite, W. Wu, H.W. Mankouri, H. Shen, Y. Liu, I.D. Hickson, Replication stress activates DNA repair synthesis in mitosis, Nature, 528 (2015) 286-290.
  12. L. Ren, L. Chen, W. Wu, L. Garribba, H. Tian, Z. Liu, I. Vogel, C. Li, I.D. Hickson, Y. Liu, Potential biomarkers of DNA replication stress in cancer, Oncotarget, 8 (2017) 36996-37008.
  13. V.A. Bjerregaard, L. Garribba, C.T. McMurray, I.D. Hickson, Y. Liu, Folate deficiency drives mitotic missegregation of the human FRAXA locus, Proc Natl Acad Sci U S A, (2018).
  14. S. Bursomanno, P. Beli, A.M. Khan, S. Minocherhomji, S.A. Wagner, S. Bekker-Jensen, N. Mailand, C. Choudhary, I.D. Hickson, Y. Liu, Proteome-wide analysis of SUMO2 targets in response to pathological DNA replication stress in human cells, DNA Repair (Amst), 25 (2015) 84-96.
  15. S. Bursomanno, J.F. McGouran, B.M. Kessler, I.D. Hickson, Y. Liu, Regulation of SUMO2 target proteins by the proteasome in human cells exposed to replication stress, J Proteome Res, 14 (2015) 1687-1699.