UCL Cancer Institute

Genome Organization and Function


Group Leader: Dr Suzana Hadjur




Despite the vast amount of information that the sequencing of our genome has brought, our nucleus is still a mysterious place. The mammalian genome is estimated to contain 22,000 genes which reside on 46 chromosomes. In order to contain our genome within nuclear space, DNA molecules must undergo many levels of organization resulting in discrete nuclear ‘environments’. Moreover, this organization must be dynamic and responsive as cells go about the regular business of producing proteins, responding to their environment and dividing. How this complex organization is orchestrated is an important question in genome biology.







 

Group Members

  • Dr Sevil Sofueva
  • Dr Wen-Ching Chan
  • Matteo Vietri Rudan, PhD student
  • Dimitra Georgopoulos, PhD student


Collaborators


 



All cells contain the same DNA content, however the activity of particular genes changes during development leading to specialized cells and tissues. Understanding how gene activity is controlled lies at the heart of any exploration of development. Growing evidence indicates that the spatial organization of the genome, ie. the three-dimensional configuration and localization of genes within the nucleus, plays an important role in the systems that control gene activity.

We are interested in understanding the fundamental principles behind genome organization in space and time, the proteins involved in establishing and maintaining this organization and how it changes during cellular development and disease. We use cellular biology and high-resolution molecular approaches to gain insight into how the architectural organisation of the nucleus and regulation of gene activity are functionally linked in mammalian cells.


Research

Protein-DNA interactions are the basis for most levels of chromatin organization and many proteins which have structural functions act by binding to specific genomic regions. We focus our current studies on the role of Cohesin proteins. Cohesin proteins are integral components of chromosome structure and are essential for ensuring daughter chromatids are paired immediately after replication and prior to separation at mitosis. In addition to this, it is increasingly clear that cohesin proteins participate in multiple mechanisms necessary for proper cellular behaviour which include interphase genome organization, control of gene expression and development.

The DNA binding protein CTCF has long been recognized as playing a central role in the organization of chromatin, regulating the ability of DNA elements involved in gene expression to communicate. Together with supporting work from other groups, our observations linked CTCF with cohesins and provided an explanation for how a protein complex that mediates sister chromatin cohesion is involved in regulating gene expression. This now leads to new questions about how cohesin-based chromatin topology influences gene activity.

Insight into how cohesin proteins work will lead to a better understanding of many aspects of chromosome biology as well as a deeper knowledge of developmental genetic diseases such as Roberts and Cornelia de Lange syndromes (caused by mutations in cohesin subunits) and cancer, where gene deregulation and anueploidy play a critical role.


CTCF

Fig 1. CTCF binding to cell-specific cohesin sites is directed by DNA methylation.
Shared and cell-specific cohesin sites (top panel) were assessed by ChIP for CTCF binding in B3 (dark bars) and VL3 (light bars) cells (middle panel). Methylation of CpG dinucleotides was analyzed by bisulphite sequencing of B3 and VL3 DNA (bottom panel). Methylated CpG dinucleotides are indicated by filled circles and unmethylated CpG dinucleotides by open circles. Arrows mark the position of predicted CTCF-binding sites.

.


Cohesins

Fig 2. Cohesins are required for long-range interactions at IFNG.
a) 3C analysis using a fragment containing the IFNG gene as a bait in Th1 cells (red lines) or Th1 cells treated with siRNA oligos to Rad21 (blue lines). Evidence for loss of Rad21 protein upon siRNA treatment (representative W.blot panels to the right of graph). b) ChIP analysis in Th1 cells (red bars) after Rad21 RNAi treatment (blue bars) reveals loss of Rad21 protein bound to chromatin.

.


Cohesins

Fig 3. Cohesin-dependant chromatin loops throughout the cell cycle.
G1, G2 and M nuclei are depicted as having 5 chromosomes for simplicity (blue, orange, grey, red and purple lines). Chromosome territories (CT) are depicted in G1 and G2 nuclei where a degree of intermingling is shown between CTs (loops extending from CT globules). The nucleolus has been omitted for simplicity. G2 nuclei are swollen in size and grey shadows represent replicated and cohesed chromosomes. Fully condensed and aligned chromosomes at the metaphase plate are shown in M nucleus. NPC, nuclear pore complexes; LAD, lamin-associated chromatin domains; E, enhancer element; red or orange ring, cohesin; shown only as a single green circle for ease, CTCF; grey circle, mediator or other transcription factors. a) Allele-specific loops. H19 and Igf2 are reciprocally imprinted with maternal expression of H19 and paternal expression of Igf2. Loops on the maternal allele restrict access of the Igf2 promoter to enhancers downstream of H19. b) Insulator loops partition chromatin into independently regulated domains (yellow genes and white genes). Such loops have been shown to exist at the β-globin locus and the apolipoprotein cluster. c) Interactions between a gene promoter and a distal enhancer element, either mediated by CTCF/cohesin (IFNG) or by Mediator/cohesin (Nanog). d) Multiple loops resembling a rosette structure function to gather replication origins (depicted in black), possibly tethered to a nuclear matrix. e) In G2, cohesin mediates chromatin interactions necessary for both sister chromatid cohesion (red rings) of replicated chromosomes (depicted in two shades of blue) and gene regulation (orange rings). f) Early in mitosis, cohesin dissociates from chromosome arms, while centromeric cohesin is protected and remains stable.

.


 

Selected Publications


Sofueva S and Hadjur S. (2011) Cohesin mediated chromatin interactions – into the third dimension of gene regulation. Briefings in Functional Genomics. Brief Funct Genomics. 2012 Jan 25. Pubmed

Hadjur S, Williams LM, Ryan NK, Cobb BS, Sexton T, Fraser P, Fisher AG, Merkenschlager M. Cohesins form chromosomal cis-interactions at the developmentally regulated IFNG locus. Nature. 2009 Jul 16;460(7253):410-3. Pubmed

Bowers SR, Mirabella F, Calero-Nieto FJ, Valeaux S, Hadjur S, Baxter EW, Merkenschlager M, Cockerill PN. A conserved insulator that recruits CTCF and cohesin exists between the closely related but divergently regulated interleukin-3 and granulocyte-macrophage colony-stimulating factor genes. Mol Cell Biol. 2009 Apr;29(7):1682-93. Pubmed

Parelho V*, Hadjur S*, Spivakov M, Leleu M, Sauer S, Gregson HC, Jarmuz A, Canzonetta C, Webster Z, Nesterova T, Cobb BS, Yokomori K, Dillon N, Aragon L, Fisher AG, Merkenschlager M. (2008) Cohesins functionally associate with CTCF on mammalian chromosome arms. Cell. 2008 Feb 8;132(3):422-33. Pubmed
*Equal contribution

McMahon KA, Hiew SY, Hadjur S, Veiga-Fernandes H, Menzel U, Price AJ, Kioussis D, Williams O, Brady HJ. (2007) Mll has a critical role in fetal and adult hematopoietic stem cell self-renewal. Cell Stem Cell. 2007 Sep 13;1(3):338-45. Pubmed