Proximity in chromatin : opportunities for innovations

Mammalian chromosomes extensively communicate with each other via long-range chromatin interactions. These interactions are mostly mediated by proteins, which work as teams to control genes in the cells. These interactions could also help to unravel the mechanisms of diseases such as cancer, from new perspectives. The packaging of the chromatin fiber and how it relates to epigenetic marks that regulate its accessibility to govern lineage-specific gene expression repertoires is currently the focus of immense efforts worldwide. Moreover, how chromosomes are hierarchically folded and how they relate to each other as well as to structural hallmarks of the nucleus is a largely unchartered territory in large cell populations not to mention in individual cells.

This thesis has an emphasis on the analysis of pivotal chromatin features of single cells. Thus, interactions between a genome organizer termed CTCF and a factor involved in DNA repair, PARP1, could be demonstrated using the ISPLA technique. Such interactions likely underlie the formation of chromatin networks. Next, novel strategies/techniques were developed to visualize chromosomal structures and 3D networks by scoring for chromatin proximities within individual cells.

One strategy included a novel method termed Chromatin In Situ Proximity (ChrISP) to visualize and identify proximities between chromatin fibers and other structural hallmarks in single cells at a resolution < 170 Å beyond that of the light microscope. Thus, large-scale changes in conformations of a single human chromosome upon the administration of reprogramming cues could be visualized.

Finally, this innovation was further developed to explore differences in proximities of chromatin fibers that organize chromosome territories. The novel design, termed “rainbow ChrISP” translates physical distances in 3D, between chromatin fibres into different colors visualized with conventional microscope. This technique produced new insights into chromosome conformations and their regulation to enhance our understanding of their governing principles in single cells during development and disease.

List of scientific papers

I. Dawn Farrar, Sushma Rai, Igor Chernukhin, Maja Jagodic, Yoko Ito, Samer Yammine, Rolf Ohlsson, Adele Murrell, and Elena Klenova (2010). Mutational Analysis of the Poly(ADP-Ribosyl)ation Sites of the Transcription Factor CTCF Provides an Insight into the Mechanism of Its Regulation by Poly(ADP-Ribosyl)ation. Molecular and Cellular Biology. 30(5), 1199-1216.
https://doi.org/10.1128/MCB.00827-09.

II. Xingqi Chen, Chengxi Shi, Samer Yammine, Anita Göndör, Daniel Rönnlund, Alejandro Fernandez-Woodbridge, Noriyuki Sumida, Jerker Widengren and Rolf Ohlsson. (2014). Chromatin In Situ Proximity (ChrISP): Two-parameter, single-cell analysis of chromatin proximities at a high resolution. BioTechniques. [Accepted]
https://pubmed.ncbi.nlm.nih.gov/24641475

III. Xingqi Chen, Samer Yammine, Chengxi Shi, Anita Göndör, Mariliis Tark-Dame, Noriyuki Sumida and Rolf Ohlsson. The H3K9me2/3 mark provides spatial identity and protects against large scale reorganization of a human chromosome. [Manuscript]

IV. Samer Yammine and Rolf Ohlsson. Rainbow Chromatin In Situ Proximity analysis of intra-chromosomal topologies in single interphase cells. [Manuscript]