3), in order to further study the intrinsic (between alleles) and extrinsic (cell to cell) variation in replication and to further investigate the relation between cell-to-cell heterogeneity of replication and the cell-to-cell heterogeneity in gene transcription and chromatin organization. Moreover, we have developed new genome-wide approaches to study replication program at single molecule/cell resolution (Fig. The pseudo bulk RT computed from the scRT and the population RT have a very high correlation (Pearson correlation R=0.9), and the cell-to-cell heterogeneity of replication within a cell population can be further investigated. (b) Heatmap shows the single cell replication timing (scRT) profiles based on single cell copy number variation (scCNV) detection. (a) Map early initiation events in human cells by a high-throughput single-molecule approach, called Optical Replication Mapping (ORM) that combines the Bionano Genomics approach to mapping long individual DNA molecules (green labels) with in vivo fluorescent nucleotide pulse-labeling (red) (Wang 2020). We further used these unique data to study the replication-transcription conflict and its link to genome instability.įigure 3: Study human replication program at single molecule/cell resolution. The quality and novelty of the data, leads to new insights into the replication landscape of the human genome and to further unravel the links between replication, gene expression, epigenetic modification and 3D genome organization in normal and cancer cells. A transition from rightward- to leftward-moving forks occurs when crossing a replication origin position, leading to an upward transition in the RFD profile (Fig. This allows us to determine the fractions of rightward- (R) and leftward- (L) moving forks at each locus, and to construct the complete profiles of replication fork directionality (RFD = R-L) along the genome. More recently, we have developed a new method to study the replication program based on the deep sequencing of Okazaki fragments (OK-Seq). Replication initiation poor regions nested in large gene body leads to under-replication upon fork slowing and causes CFS instability (Brison 2019). The initiation and termination zones are highlighted by orange and purple boxes. 2).įigure 2: OK-Seq, Repli-Seq (for Non-Treated and Aph treated cells), and GRO-Seq profiles are shown along regions around the indicated genes hosting Common Fragile Sites (CFSs) fine-mapped by FISH (black bars) in lymphocytes. We are now applying Repli-Seq technique to analyze the replication dynamics from cells upon replication stress to study how deregulation of the replication program challenges genome stability, in particular, common fragile site activity (Fig. By evolutionary analyses, we have also established that replication is a major process driving genome mutational landscape in normal and cancer cells. Studies of these profiles from different human cell types have allowed us to reveal that the genome is organized in megabase replication domains associated with higher order chromatin structural units. In collaboration with experimental biologists, we have developed a method (Repli-Seq) and generated one of the first high-resolution replication timing profiles of the human genome (Fig. The ascending and descending segments on the RFD profiles indicate the locations of replication initiation and termination zones, respectively. Figure 1: Spatio-temporal replication program of the human genome revealed by Ok-Seq (Replication Fork Directionality, RFD) and Repli-Seq (Replication Timing).
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |