BeNeLux Bioinformatics Conference – Antwerp, December 7-8 <strong>2015</strong> Abstract ID: P Poster 10th Benelux Bioinformatics Conference <strong>bbc</strong> <strong>2015</strong> P70. THE IMPACT OF HMGA PROTEINS ON REPLICATION ORIGINS DISTRIBUTION A. Zouaoui 1 , M. Kahli 2 , E. Besnard 3 , R. Desprat 1 , N. Kirsten 4 , P. Ben-sadoun 1 & J.M. Lemaitre 1 . Institute for Regenerative Medicine and Biotherapy, France 1 ; Institut de Biologie de l’École Normale Supérieure (ENS), France 2 ; The Gladstone Institutes, University of California San Francisco (UCSF), United States 3 ; Helmholtz Zentrum München, Research Unit Gene Vectors, Munich, Germany 4 . Proliferative cells can have an irreversible stop in the cell cycle that is called cellular senescence which can induct the development of cancer and ageing. Senescence is characterized by the development of Dense Heterochromatic Foci (SAHF) and the decline of the DNA replication. High-Mobility Group A proteins promote SAHF formation, a proliferative stop and stabilize senescence when overexpressed. In a cell, DNA replication is regulated on several genomics sites called replication origin (« Oris »). Prereplication proteic complex is required for DNA replication to occur. In the pre-replication complex, the ORC1 protein is involved in recognition of the origin of replication. DNA autoradiography of eukaryote cells allowed to find that human replication origins are bidirectional and spaced at 20-400kb intervals (Huberman and Riggs, 1968). At each origin, replication forks are formed and new short nascent strand are synthetized. A popular method to map replication origins is the purification of Short Nascent Strand (SNS). Several laboratories have identified up to 50 000 origins using microarray and sequencing techniques. Our laboratory has developed an origin mapping method divided in four cell type: IMR90, H9, iPSC and HeLa (Besnard et al., 2012). The Short Nascent Strand was isolated, sequenced and analyzed. 250 000 origin peaks have been identified with a peak detection tool named SoleSearch (Blahnik KR, Dou L, O’Geen H, et al. 2010). The objective is to find the most sensitive method to analyze the origin distribution in proliferative and senescent cells to observe if senescence has an impact on the origin distribution. The implication of HMGA proteins on the DNA replication is investigated. Two new methods are in development to analyze the replication origin with two more sensitive tools. In the first method, we search origin peaks with Macs2 tool (Zhang et al., 2008) which uses a new statistic and algorithm model. In a second time, origin enrichment is observed with Homer tool (Heinz S et al., 2010). Two methods are currently in development to identify the replication origin site by Illumina GaII sequencing of short nascent strand. Human SNS-seq reads of 36bp were mapped to human genome build GRCH38 with BWA tool (ref). Origin peaks were called by MACS2 and origin enrichment by Homer. To compare the two methods, active origins in HeLa cells were detected with each method. Correlation between ORC1 peaks and origins identified is calculated to choose the most sensitive method. The impact of pre-senecence is observed in comparing origins distribution observed in proliferative and senescent cells. Origins distribution is compared before and after induction of HMGA proteins to investigate the implication of these proteins on the DNA replication during senescence. REFERENCES Besnard et al. Best practices for mapping replication origins in eukaryotic chromosomes. Current Protoc Cell Biol. 2014 Sep 2; 64:22.18.1-22.18.13 Besnard et al. Unraveling cell type-specific and reprogrammable human replication origin signatures associated with G-quadruplex consensus motifs. Nat Struct Mol Biol. 2012 Aug; 19, 837-44 Blahnik KR, Dou L, O’Geen H, et al. Sole-Search: an integrated analysis program for peak detection and functional annotation using ChIP-seq data. Nucleic Acids Res. 2010; 38:e13 Fu H et al. Mapping replication origin sequences in eukaryotic chromosomes. Curr Protoc Cell Biol. 2014 Dec 1; 65:22.20.1- 22.20.17 Heinz S, Benner C, Spann N, Bertolino E et al. Simple Combinations of Lineage-Determining Transcription Factors Prime cis-Regulatory Elements Required for Macrophage and B Cell Identities. Mol Cell 2010 May 28; 38, 576-589 Hubberman JA et al. On the mechanism of DNA replication in mammalian chromosomes. J Mol Biol 1968 Mar 14; 32, 327-41 Zhang et al. Model-based Analysis of ChIP-Seq (MACS). Genome Biol (2008) 9 pp. R13 114
10th Benelux Bioinformatics Conference <strong>bbc</strong> <strong>2015</strong> <strong>bbc</strong> <strong>2015</strong> December 7 - 8, <strong>2015</strong> Antwerp, Belgium www.<strong>bbc</strong><strong>2015</strong>.be 115
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